SRX5395624 GSM3613463 SRP186374 SRS4382562 GSM3613463 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613463 GSM3613463 GEO Accession GSM3613463 SRX5395625 GSM3613464 SRP186374 SRS4382563 GSM3613464 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613464 GSM3613464 GEO Accession GSM3613464 SRX5395626 GSM3613465 SRP186374 SRS4382564 GSM3613465 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613465 GSM3613465 GEO Accession GSM3613465 SRX5395627 GSM3613466 SRP186374 SRS4382565 GSM3613466 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613466 GSM3613466 GEO Accession GSM3613466 SRX5395628 GSM3613467 SRP186374 SRS4382566 GSM3613467 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613467 GSM3613467 GEO Accession GSM3613467 SRX5395629 GSM3613468 SRP186374 SRS4382567 GSM3613468 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613468 GSM3613468 GEO Accession GSM3613468 SRX5395630 GSM3613469 SRP186374 SRS4382568 GSM3613469 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613469 GSM3613469 GEO Accession GSM3613469 SRX5395631 GSM3613470 SRP186374 SRS4382569 GSM3613470 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 303613470 GSM3613470 GEO Accession GSM3613470 SRX5395632 GSM3613471 SRP186374 SRS4382570 GSM3613471 RIP-Seq TRANSCRIPTOMIC other Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613471 GSM3613471 GEO Accession GSM3613471 SRX5395633 GSM3613472 SRP186374 SRS4382571 GSM3613472 RIP-Seq TRANSCRIPTOMIC other Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613472 GSM3613472 GEO Accession GSM3613472 SRX5395634 GSM3613473 SRP186374 SRS4382572 GSM3613473 RIP-Seq TRANSCRIPTOMIC other Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613473 GSM3613473 GEO Accession GSM3613473 SRX5395635 GSM3613474 SRP186374 SRS4382573 GSM3613474 RIP-Seq TRANSCRIPTOMIC other Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613474 GSM3613474 GEO Accession GSM3613474 SRX5395636 GSM3613475 SRP186374 SRS4382574 GSM3613475 ncRNA-Seq TRANSCRIPTOMIC size fractionation Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613475 GSM3613475 GEO Accession GSM3613475 SRX5395637 GSM3613476 SRP186374 SRS4382575 GSM3613476 ncRNA-Seq TRANSCRIPTOMIC size fractionation Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613476 GSM3613476 GEO Accession GSM3613476 SRX5395638 GSM3613477 SRP186374 SRS4382576 GSM3613477 ncRNA-Seq TRANSCRIPTOMIC size fractionation Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613477 GSM3613477 GEO Accession GSM3613477 SRX5395639 GSM3613478 SRP186374 SRS4382577 GSM3613478 ncRNA-Seq TRANSCRIPTOMIC size fractionation Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. NextSeq 500 gds 303613478 GSM3613478 GEO Accession GSM3613478 SRX9251232 GSM4819873 SRP186374 PRJNA523364 SRS7483691 GSM4819873 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 304819873 GSM4819873 GEO Accession GSM4819873 SRX9251233 GSM4819874 SRP186374 PRJNA523364 SRS7483692 GSM4819874 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 304819874 GSM4819874 GEO Accession GSM4819874 SRX9251234 GSM4819875 SRP186374 PRJNA523364 SRS7483693 GSM4819875 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 304819875 GSM4819875 GEO Accession GSM4819875 SRX9251235 GSM4819876 SRP186374 PRJNA523364 SRS7483694 GSM4819876 RNA-Seq TRANSCRIPTOMIC cDNA Total RNA was extracted by TRIZOL reagent. For RIP-seq, N2a cells cultured in a 150 mm plate were washed twice with cold PBS followed by UV irradiation at 400mJ/cm2 under a HL-2000Hybrilinker. Scrapped cells were harvested by centrifugation at 500 g for 5 min at 4°C. Crosslinked cells were lysed in 1 ml lysis buffer (20mM Tris-Hcl pH 7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 0.1% SDS, 0.1% deoxycholate, 100U/ml RNasin Ribonuclease Inhibitors (Promega), 1 u/ml EDTA-free complete protease inhibitor cocktail (Roche)) on ice for 30 min. Lysates were centrifuged at 12,000 g at 4°C for 15 min to remove cell debris. An input of collected supernatant was set aside for subsequent analysis. Each sample was then divided and incubated with 5 μg of either anti-TDP-43 antibody (Proteintech) or rabbit IgG (Millipore) on a rotator overnight at 4°C. Complexes were pulled down by incubation with protein G Dynabeads on a rotator for 2 hours at 4°C. Beads were washed twice with lysis buffer, twice with wash buffer (add 300 mM NaCl), and boiled to reverse UV cross-linking. Co-precipitated RNAs were extracted from beads and used for construction of small RNA library or analysis by quantitative RT-PCR (qRT-PCR) using specific primers.To isolate cellular insoluble aggregates, two 10-cm dish N2a cells were extracted sequentially, as previously described (Guo et al., 2011; Hu et al., 2017). Briefly, cells were harvested, lysed in the moderate RIPA lysis buffer (50mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, EDTA-free complete protease inhibitor cocktail (Roche), 100U/ml RNasin Ribonuclease Inhibitors (Promega)) on ice for 30 min, and centrifuged at 17000 g at 4°C for 30 min. The bottom cell debris (i.e. nuclear, mitochondria and stress granules) were depleted. The resultant supernatant fractions were incubated with 1% Sarkosyl for 30 min at room temperature with agitation, and then the detergent insoluble proteins were obtained by ultracentrifugation of the supernatants at 41600 rpm at 4°C for 30 min and washed twice by PBS buffer. The whole cell lysates and the detergent-insoluble pellets were separately collected, and the isolated protein and RNA were used for Western blotting analysis and small RNA-seq library construction.The small RNA library was constructed using sensitivity-improved method as described (Yang et al., 2016) or modifications. Briefly, isolated total RNA (~1 μg) was mixed with 25 pmol 3′ adapter, 1 × T4 ligase reaction buffer, 33% polyethylene glycol (molecular weight 8000), 40 U RNL2tr K227Q (NEB), and 20 U RiboLock RNase Inhibitor (Thermo Fisher). The samples were incubated at 22°C for 2 hours, then at 16°C for 1 hour, followed by addition of 1 μl of 10 μM RT primer before heat inactivation at 75°C for 5 min. 200 pmol of the 5′ adapter was added for ligation by T4 RNA ligase 1 (NEB) and incubated at 22°C for 1 hour. Linker-ligated RNAs were reverse transcribed with Superscript III (Invitrogen) at 50°C for 30 min, 55°C for 30 min, then at 95°C for 5 min and PCR-amplified with Q5 High-Fidelity 2× Master (NEB). The product was fractionated on a 4% agarose gel and extracted by QIAEX II Gel Extraction Kit (QIAGEN). The recovered DNA was quantified for deep sequencing. Illumina HiSeq 2000 gds 304819876 GSM4819876 GEO Accession GSM4819876