SRP435437 PRJNA965818 GSE231436 Multi-omic study of genome-edited human colonoids reveal patterns of microRNA regulation specific to different colorectal cancer mutation profiles Colorectal cancer (CRC) is the third most diagnosed cancer and the second leading cause of cancer-related death worldwide Unique combinations of mutations can affect responsiveness to specific therapeutics. This can be, at least in part, due to mutation-specific alterations in gene regulatory mechanisms. To better understand how unique combinations of mutations affect gene regulation, we generated small RNA-seq, length extension chromatin run-on-seq (leChRO-seq), and RNA-seq data from APC (A-mutant), APC/KRAS (AK-mutant), APC/KRAS/TP53 mutant (AKP-mutant), and iGFP control human colonic organoids. From these analyses, we found that our colonic organoid models demonstrate mutation-specific patterns of miRNA transcription, miRNA expression and transcriptional regulatory element actvitiy. Furthermore, we defined 10 patterns of miRNA expression across organoid models. We highlighted one group of miRNAs that exhibited a unique downregulation of expression in AKP-mutant organoids (including miR-34a-5p and miR-10a-5p). Analysis of miRNA transcription revealed that most changes in miRNA expression are correlated with changes in miRNA transcription. Analysis of leChRO-seq data revealed that transcriptional regulatory elements upregulated in AKP-mutant colonic organoids have an enrichment of predicted binding sites for oncogenic transcription factors. Overall design: Genetically modfieid colonic organoid (termed colonoid) models were generated using a dox-inducible CRISPR system. smRNA-seq data was generated for APC+dox mutant (A-mutant; n=3), APC/KRAS+dox mutant (AK-mutant; n=4), APC/KRAS/TP53+dox mutant (AKP-mutant; n=2), iGFP+dox (iGFP_dox; n=4), iGFP-dox (iGFP_noDox; n=4). Length extension chromatin run-on sequencing (leChRO-seq) data was generated for A-mutant (n=2), AK-mutant (n=2), AKP-mutant (n=2), iGFP_dox (n=2), and iGFP_noDox (n=2). RNA-seq data was generated for iGFP_dox (n=4) and AKP-mutant (n=3). GSE231436