Chemical genetic approaches to study chromatin complexes - the epigenome comprises a critical layer for controlling gene expression and genome function

Cancer mutations often alter the function of chromatin complexes, leading to aberrant epigenomic landscapes frequently observed in tumor cells. Determining the mechanisms controlling chromatin complexes and their interactions will advance our understanding of epigenomic processes, how they are disrupted in cancer, and how they can be pharmacologically modulated for drug discovery. Consequently, our central goals are to elucidate the mechanisms of chromatin complexes and test their promise as therapeutic targets. In this pursuit, this application investigates lysine-specific histone demethylase-1 (LSD1), a transcriptional corepressor that is a drug target for oncology. LSD1 forms complexes with various corepressors and transcription factors (TF), including GFI1B, which are critically involved in development and implicated across various tumor types. Using drug-resistance alleles obtained from a chemical suppressor screen, our prior work showed that LSD1 active site inhibitors exert their anti-proliferative effects by disrupting the LSD1-GFI1B complex, revising prior models of drug mechanism of action. Notably, GFI1B is frequently overexpressed by enhancer hijacking mutations in group 3 and 4 medulloblastoma (MB), and LSD1 inhibitors are effective in GFI1B-driven MB mouse models. Intriguingly, the E3 ubiquitin ligase KBTBD4 is also frequently mutated in group 3/4 MB and was recently reported to mediate degradation of corest, LSD1'S obligate complex partner. These observations suggest a possible mechanistic connection between GFI1B and KBTBD4 in group 3/4 MB, mediated through LSD1-corest. However, the molecular interactions and interplay between LSD1-corest, GFI1B, and KBTBD4 remain unclear and present a major GAP in our understanding. To address these gaps, the first specific aim investigates the structure, dynamics, and interactions of the LSD1-GFI1B complex through a multidisciplinary approach, with the goal of revealing an unprecedented view into a chromatin regulator-TF complex. The second aim seeks to elucidate the mechanism of small molecules that degrade LSD1-corest by potentiating KBTBD4 activity, providing critical insight into strategies to target LSD1 complexes through new emerging modalities. The last aim studies how KBTBD4 MB mutations promote LSD1-corest degradation and their downstream consequences on LSD1- GFI1B and the MB cancer epigenome. Across these aims, the mechanisms and interactions of LSD1 complexes will be explored by using innovative chemical genomic approaches that leverage drug suppressor alleles with cell, molecular, and structural biology. IT is expected that the findings from these studies will illuminate biochemical principles governing the function and interactions of chromatin complexes and advance strategies to pharmacologically target them for therapeutic applications.