Multi-Omics Elucidation of KDM5C, KDM6A, and KMT2B Roles in Cancer Epigenetic Dysregulation and Transcriptional Reprogramming [ATAC-seq]

Histone-modifying enzymes (HMEs) are critical regulators of tumorigenesis through epigenetic reprogramming. While mutations in HMEs are recognized drivers of cancer epigenome dysregulation, systematic comparative analyses of their mutational impacts and functional divergence across malignancies remain underexplored. This study addresses this gap by investigating three HMEs—KDM5C (H3K4me3 demethylase), KMT2B (H3K4me3 methyltransferase), and KDM6A (H3K27me3 demethylase)—frequently mutated in diverse cancers. Using CRISPR/Cas9 knockout cell lines, we performed integrated multi-omics profiling encompassing genome-wide chromatin accessibility, transcriptomics, and chromatin-bound proteomics. Contrary to expectations that KMT2B loss (H3K4me3 depletion) and KDM5C loss (H3K4me3 accumulation) would induce opposing transcriptional programs, or that KDM6A deficiency (H3K27me3 accumulation) would exhibit distinct regulatory effects, our analyses revealed striking discrepancies between transcriptional outputs and chromatin-associated proteomic states across all three knockouts. Notably, each HME knockout elicited distinct transcriptional regulatory patterns, challenging conventional assumptions about their antagonistic or synergistic functions. These findings highlight context-dependent functional hierarchies among HMEs and underscore the necessity of multi-dimensional profiling to resolve epigenetic regulatory complexity. Our work advances the mechanistic understanding of cancer epigenetics and provides a framework for elucidating tumorigenic vulnerabilities linked to HME dysregulation.