Chemical Genetic Approaches to Study Chromatin Complexes

表观基因组包括控制基因表达和基因组功能的关键层。癌 突变经常改变染色质复合体的功能，导致经常出现异常的表观基因组图谱。 在肿瘤细胞中观察到。确定控制染色质复合体及其相互作用的机制将 促进我们对表观基因组过程的理解，它们在癌症中是如何被破坏的，以及它们是如何被破坏的 为药物发现做好了药理调整。因此，我们的中心目标是阐明 研究染色质复合体的作用机制，并测试其作为治疗靶点的前景。在这一追求中，这个应用程序 研究赖氨酸特异的组蛋白去甲基酶-1(LSD1)，这是一种转录辅助抑制物，是药物靶点 肿瘤学。LSD1与多种辅阻遏子和转录因子(Tf)形成复合体，包括GFI1B， 它们与肿瘤的发生和发展密切相关，并与各种肿瘤类型有关。利用抗药性 从化学抑制物筛选获得的等位基因，我们先前的工作表明LSD1活性位点抑制物对 它们通过破坏LSD1-GFI1B复合体，修正先前的药物机制模型而发挥抗增殖作用 行动的一部分。值得注意的是，GFI1B在组3和组4中经常由于增强子劫持突变而过度表达 髓母细胞瘤(MB)和LSD1抑制剂在GFI1B驱动的MB小鼠模型中有效。有趣的是， E3泛素连接酶KBTBD4在3/4MB组中也经常发生突变，最近有报道称 核心退化，LSD1 S包办复杂搭档这些观察表明一种可能的机械论 3/4MB组GFI1B与KBTBD4之间的连接通过LSD1-corest介导。然而， LSD1-COREST、GFI1B和KBTBD4之间的分子相互作用和相互作用仍不清楚和存在 这是我们在理解上的一个重大差距。为了解决这些差距，第一个具体目标是调查结构， 通过多学科方法研究LSD1-GFI1B复合体的动力学和相互作用，目标是 揭示了染色质调节因子-转铁蛋白复合体的前所未有的视角。第二个目的是试图澄清 小分子通过增强KBTBD4活性降解LSD1-corest的机制，提供关键的 洞察通过新的新兴模式瞄准LSD1复合体的战略。最后一个目标是研究如何 KBTBD4 MB突变促进LSD1核心降解及其对LSD1下游的影响 GFI1B和MB癌症表观基因组。跨越这些目标，LSD1复合体的机制和相互作用 将通过使用创新的化学基因组方法来探索，这些方法利用药物抑制基因与 细胞、分子和结构生物学。预计这些研究的结果将阐明 染色质复合体功能和相互作用的生化原理及发展策略 将它们作为治疗应用的药理靶点。

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.