SRX22251811 GSM7864602_r1 SRP468872 PRJNA1032006 SRS19305474 GSM7864602 GSM7864602 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251812 GSM7864603_r1 SRP468872 PRJNA1032006 SRS19305475 GSM7864603 GSM7864603 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251813 GSM7864604_r1 SRP468872 PRJNA1032006 SRS19305476 GSM7864604 GSM7864604 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251814 GSM7864605_r1 SRP468872 PRJNA1032006 SRS19305477 GSM7864605 GSM7864605 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251815 GSM7864608_r1 SRP468872 PRJNA1032006 SRS19305478 GSM7864608 GSM7864608 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251816 GSM7864609_r1 SRP468872 PRJNA1032006 SRS19305479 GSM7864609 GSM7864609 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251817 GSM7864610_r1 SRP468872 PRJNA1032006 SRS19305480 GSM7864610 GSM7864610 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251818 GSM7864611_r1 SRP468872 PRJNA1032006 SRS19305481 GSM7864611 GSM7864611 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251819 GSM7864612_r1 SRP468872 PRJNA1032006 SRS19305482 GSM7864612 GSM7864612 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251820 GSM7864613_r1 SRP468872 PRJNA1032006 SRS19305483 GSM7864613 GSM7864613 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251821 GSM7864606_r1 SRP468872 PRJNA1032006 SRS19305484 GSM7864606 GSM7864606 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251822 GSM7864607_r1 SRP468872 PRJNA1032006 SRS19305485 GSM7864607 GSM7864607 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251823 GSM7864634_r1 SRP468872 PRJNA1032006 SRS19305486 GSM7864634 GSM7864634 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251824 GSM7864614_r1 SRP468872 PRJNA1032006 SRS19305487 GSM7864614 GSM7864614 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251825 GSM7864615_r1 SRP468872 PRJNA1032006 SRS19305488 GSM7864615 GSM7864615 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251826 GSM7864616_r1 SRP468872 PRJNA1032006 SRS19305489 GSM7864616 GSM7864616 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251827 GSM7864617_r1 SRP468872 PRJNA1032006 SRS19305490 GSM7864617 GSM7864617 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251828 GSM7864618_r1 SRP468872 PRJNA1032006 SRS19305491 GSM7864618 GSM7864618 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251829 GSM7864619_r1 SRP468872 PRJNA1032006 SRS19305492 GSM7864619 GSM7864619 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251830 GSM7864620_r1 SRP468872 PRJNA1032006 SRS19305493 GSM7864620 GSM7864620 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251831 GSM7864621_r1 SRP468872 PRJNA1032006 SRS19305494 GSM7864621 GSM7864621 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251832 GSM7864641_r1 SRP468872 PRJNA1032006 SRS19305495 GSM7864641 GSM7864641 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251833 GSM7864648_r1 SRP468872 PRJNA1032006 SRS19305496 GSM7864648 GSM7864648 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251834 GSM7864622_r1 SRP468872 PRJNA1032006 SRS19305497 GSM7864622 GSM7864622 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251835 GSM7864623_r1 SRP468872 PRJNA1032006 SRS19305498 GSM7864623 GSM7864623 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251836 GSM7864624_r1 SRP468872 PRJNA1032006 SRS19305499 GSM7864624 GSM7864624 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251837 GSM7864625_r1 SRP468872 PRJNA1032006 SRS19305500 GSM7864625 GSM7864625 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251838 GSM7864626_r1 SRP468872 PRJNA1032006 SRS19305501 GSM7864626 GSM7864626 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251839 GSM7864627_r1 SRP468872 PRJNA1032006 SRS19305502 GSM7864627 GSM7864627 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251840 GSM7864628_r1 SRP468872 PRJNA1032006 SRS19305503 GSM7864628 GSM7864628 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251841 GSM7864629_r1 SRP468872 PRJNA1032006 SRS19305504 GSM7864629 GSM7864629 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251842 GSM7864630_r1 SRP468872 PRJNA1032006 SRS19305505 GSM7864630 GSM7864630 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251843 GSM7864631_r1 SRP468872 PRJNA1032006 SRS19305506 GSM7864631 GSM7864631 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251844 GSM7864632_r1 SRP468872 PRJNA1032006 SRS19305507 GSM7864632 GSM7864632 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251845 GSM7864633_r1 SRP468872 PRJNA1032006 SRS19305508 GSM7864633 GSM7864633 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251846 GSM7864635_r1 SRP468872 PRJNA1032006 SRS19305509 GSM7864635 GSM7864635 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251847 GSM7864636_r1 SRP468872 PRJNA1032006 SRS19305510 GSM7864636 GSM7864636 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251848 GSM7864637_r1 SRP468872 PRJNA1032006 SRS19305511 GSM7864637 GSM7864637 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251849 GSM7864638_r1 SRP468872 PRJNA1032006 SRS19305512 GSM7864638 GSM7864638 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251850 GSM7864639_r1 SRP468872 PRJNA1032006 SRS19305514 GSM7864639 GSM7864639 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251851 GSM7864640_r1 SRP468872 PRJNA1032006 SRS19305513 GSM7864640 GSM7864640 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251852 GSM7864642_r1 SRP468872 PRJNA1032006 SRS19305515 GSM7864642 GSM7864642 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251853 GSM7864643_r1 SRP468872 PRJNA1032006 SRS19305516 GSM7864643 GSM7864643 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251854 GSM7864644_r1 SRP468872 PRJNA1032006 SRS19305517 GSM7864644 GSM7864644 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251855 GSM7864645_r1 SRP468872 PRJNA1032006 SRS19305518 GSM7864645 GSM7864645 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251856 GSM7864646_r1 SRP468872 PRJNA1032006 SRS19305519 GSM7864646 GSM7864646 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251857 GSM7864647_r1 SRP468872 PRJNA1032006 SRS19305520 GSM7864647 GSM7864647 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251858 GSM7864649_r1 SRP468872 PRJNA1032006 SRS19305521 GSM7864649 GSM7864649 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251859 GSM7864650_r1 SRP468872 PRJNA1032006 SRS19305522 GSM7864650 GSM7864650 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251860 GSM7864651_r1 SRP468872 PRJNA1032006 SRS19305523 GSM7864651 GSM7864651 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251861 GSM7864652_r1 SRP468872 PRJNA1032006 SRS19305524 GSM7864652 GSM7864652 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251862 GSM7864653_r1 SRP468872 PRJNA1032006 SRS19305525 GSM7864653 GSM7864653 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251863 GSM7864654_r1 SRP468872 PRJNA1032006 SRS19305527 GSM7864654 GSM7864654 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000 SRX22251864 GSM7864655_r1 SRP468872 PRJNA1032006 SRS19305526 GSM7864655 GSM7864655 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Nuclei were isolated from frozen tissue as previously described.10 For mouse snRNA-seq, hemicerebella of three animals per genotype (WT and SCA1 KI) per timepoint (5, 12, 18, 24, and 30 weeks) were used, for a total of 30 animals. For human post-mortem samples, pieces of cerebellar cortex were harvested from frozen tissue of ten unique individuals per diagnosis (CTRL and SCA1), for a total of 20 human samples. Briefly, frozen mouse or human tissue was gently dounce homogenized in 2mL of ice-cold Nuclei EZ Prep buffer (Sigma; NUC101-1KT) with large clearance pestle “A” and then a small clearance pestle “B” for 25 strokes each, then incubated on ice for five minutes with an additional 2mL of cold EZ Prep buffer. Nuclei were centrifuged at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of cold EZ Prep buffer, incubated on ice at five minutes, and centrifuged again at 500xg for five minutes at 4°C. Pelleted nuclei were then washed in 4mL of Nuclei Suspension Buffer (NSB) containing 1x phosphate buffered saline (PBS), 0.01% BSA, and 0.1% RNase inhibitor (Clontech/Takara; 2313B) and centrifuged at 500xg for five minutes at 4°C. Finally, nuclei were resuspended in 1mL of NSB, filtered through a 40mm cell strainer (Fisher Scientific; 22-363-547), and counted using a Countess II FL Cell Counter (Applied Biosystems; A27978). Single-nucleus suspensions were diluted to approximately 1000 nuclei per ml in NSB. Libraries were prepared from diluted single-nucleus suspensions using the 10x Genomics Chromium Single Cell 3' Reagent Kits v.3 (mouse dataset) or v.3.1 (human dataset) through the Yale Center for Genome Analysis (YCGA). Briefly, 10,000 cells per sample were added to RT Master Mix, loaded on the Single Cell A Chip, and portioned with a pool of about 750,000 barcoded gel beads to form nanoliter-scale Gel Beads-In-Emulsions (GEMs). Each gel bead has primers containing the following: an Illumina R1 sequence (read 1 sequencing primer), a 16-nucleotide barcode, a 12-nucleotide unique molecular identifier (UMI), and a 30-nucleotide poly-dT primer sequence. Upon dissolution of the Gel Beads in a GEM, the primers were released and mixed with the cell lysate and Master Mix. Incubation of the GEMs then produced barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were then used to remove leftover biochemical reagents from the post-GEM reaction mixture. Full-length, barcoded cDNA was then amplified by PCR to generate sufficient mass for library construction, and enzymatic fragmentation and size selection were used to optimize the cDNA amplicon size prior to library construction. P5, P7, a sample index, and R2 (read 2 primer sequence) were added during library construction via end repair, A-tailing, adaptor ligation, and PCR. The final libraries contained the P5 and P7 primers used in Illumina bridge amplification. Libraries were sequenced using the NovaSeq6000 at the YCGA. Sample concentrations were normalized to 1.2nM and loaded onto an Illumina NovaSeq S4 flow cell at a concentration that yielded 20k-50k passing filter clusters per cell per sample. Samples were sequenced using paired-end sequencing on an Illumina NovaSeq6000 instrument according to Illumina protocols using 10x sequencing specifications. The 8bp index was read during an additional sequencing read that automatically followed the completion of read 1. Data generated during sequencing runs were simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina was spiked into every lane at a concentration of 0.3% to monitor quality in real time. Signal intensities were converted to individual base calls during a run using the system's Real Time Analysis (RTA) software. Base calls were transferred from the machine's dedicated personal computer to the Yale high-performance computing cluster via a 1 Gigabit network mount for downstream analysis. snRNA-seq Illumina NovaSeq 6000