The COMPASS family of H3K4 methylases in development and cancer

描述(由申请人提供)： 项目概述在过去的17年里，我的实验室的研究主要集中在对Set1/MLL蛋白质家族的分子功能和生化特性的表征上。它们的嵌合体和突变与儿童白血病和其他形式的癌症有关。我们希望我们的分子研究将通过这一蛋白质家族促进我们对重排和基于突变的癌症的分子机制的理解。在这笔赠款的当前资金周期内，我的实验室已经在包括果蝇和哺乳动物细胞培养在内的多个模型系统中应用了遗传学和生物化学。我们证明了果蝇细胞具有三种Set1相关蛋白：dSet1，Trithorax(Trx)和Trithorax-Related(TRR)，所有这些蛋白都存在于能够甲基化组蛋白H3K4的指南针状成分中。哺乳动物细胞在果蝇中发现的三个亚类中的每一个都有两个代表，总共有六个COMPASS家族成员：SET1A/SET1B(与dSet1相关)；MLL1和ML2(与Trx相关)；以及ML3和MLL4(与TRR相关)。此外，考虑到许多MLL易位配对之间几乎没有序列同源性，多年来，人们一直不清楚为什么MLL易位到如此多无关基因导致白血病的发病。我们对MLL-嵌合体纯化的生化研究表明，许多MLL易位伙伴是同一大分子复合体的一部分，我们称之为超级延长复合体(SEC)。我们证明，在SEC的任何一个亚基内的MLL易位导致SEC错误地招募到MLL靶基因，并扰乱这些基因的转录检查点控制，从而触发白血病的生长。此外，我们所了解到的是，最近对癌症体细胞突变的编目发现，在血液系统恶性肿瘤和实体瘤中，MLL1-4和Set1A/B复合体的成分都存在大量突变。事实上，在不同形式的癌症中，MLL1-4和Set1A/B共同携带的突变似乎比p53更多。然而，我们对处于发育中的MLL1-4和Set1A/B家族以及它们的突变与癌症相关的原因知之甚少。鉴于在过去的17年里，我们已经在多个模型系统中开发了一套针对这些因子及其相关蛋白质的奇妙的试剂和工具，我的实验室处于非常独特的位置来定义这些因子参与癌症发病机制的分子基础，以达到靶向治疗的目的。因此，这一新的应用的目标是全面的分子和生化表征MLL1-4和Set1A-B及其复合体在基因表达和发育调控中的作用，以及它们的突变如何在人类癌症发病机制中发挥作用。此次续签申请的目标将通过三个具体目标积极实现。具体目标1集中在TRX/COMPASS家族成员(MLL1和MLL2)的分子特性的表征；它们的分子特性的鉴定和对染色质的特异性募集；以及它们的易位如何在白血病发病中起作用。具体目标2将集中在确定TRR/COMPASS家族(ML3和MLL4)在增强子单甲基化中的作用，以及增强子故障如何通过该家族成员的特定突变导致癌症在发病机制中的作用。具体目标3集中在Set1/Compass家族(Set1A和Set1B)的分子特征和结构研究，以及由这类酶实现的组蛋白H3K4三甲基化如何参与整个发育过程中基因表达的调节。我们将在多个模型系统中利用各种生化、分子和遗传工具来解决所提出的目标。这些研究应该(I)有助于我们理解COMPASS家族在调节H3K4甲基化模式中的不同作用以及它们如何调控发育和分化；以及(Ii)对于我们理解COMPASS家族内的突变如何导致癌症具有根本性的影响。这些信息有可能被证明对试图设计合理的癌症治疗方法的调查人员有帮助。

DESCRIPTION (provided by applicant): Project Summary Studies from my laboratory during the past seventeen years have focused on the characterization of the molecular functions and biochemical properties of the Set1/MLL family of proteins. Their chimeras and mutations are associated with childhood leukemia and other forms of cancers. Our hope is that our molecular studies will advance our understanding of the molecular mechanisms of rearrangement and mutation-based cancer through this family of proteins. During the current funding cycle of this grant, my laboratory has employed genetics and biochemistry in multiple model systems including Drosophila and mammalian cell cultures. We demonstrated that Drosophila cells possess three Set1-related proteins: dSet1, Trithorax (Trx), and Trithorax-related (Trr), all found within COMPASS-like compositions capable of methylating histone H3K4. Mammalian cells possess two representatives for each of the three subclasses found in Drosophila for a total of six COMPASS family members: SET1A/SET1B (related to dSet1); MLL1 and MLL2 (related to Trx); and MLL3 and MLL4 (related to Trr). Furthermore, given that there is almost no sequence homology between many of the MLL translocation partners, for many years, it was unclear why MLL translocations into so many unrelated genes result in the pathogenesis of leukemia. Our biochemical studies on the purification of the MLL-chimeras demonstrated that many of the MLL translocation partners are part of the same macromolecular complex we named the Super Elongation Complex (SEC). We demonstrated that the translocations of MLL within any of the subunits of SEC result in the misrecruitment of SEC to the MLL target genes and in the perturbation of the transcriptional checkpoint control of these genes, triggering leukemic growth. Additionally, what we have learned is that recent cataloging of somatic mutations in cancer have identified a large number of mutations in the components of the MLL1-4 and Set1A/B complexes in both hematological malignancies and solid tumors. As a matter of fact, collectively MLL1-4 and Set1A/B appear to bear more mutations in different forms of cancers than p53. However, we know very little about the MLL1-4 and Set1A/B families in development and why their mutations are associated with cancer. Given that we have developed a fantastic set of reagents and tools within the past seventeen years towards these factors and their associated proteins in multiple model systems, my laboratory is in a very unique position to define the molecular bases of these factors' involvement in cancer pathogenesis for the purpose of targeted therapeutics. Therefore, the goals of this renewal application are the full molecular and biochemical characterization of MLL1-4 and Set1A-B and their complexes in the regulation of gene expression and development and how their mutations contribute to the pathogenesis of human cancer. The goals of this renewal application will be aggressively pursued via three specific aims. Specific Aim 1 is focused on the characterization of the molecular properties of the Trx/COMPASS family members (MLL1 and MLL2); identification of their molecular properties and specific recruitment to chromatin; and how their translocations contribute to leukemic pathogenesis. Specific Aim 2 will be focused in defining the role of the Trr/COMPASS family (MLL3 and MLL4) in enhancer monomethylation and how enhancer malfunction through specific mutations of the components of this family result cancer in pathogenesis. Specific Aim 3 is focused on the molecular characterization and structural studies of the Set1/COMPASS family (Set1A and Set1B) and how histone H3K4 trimethylation implemented by this class of enzymes is involved in the regulation of gene expression throughout development. We will take advantage of a variety of biochemical, molecular, and genetic tools in multiple model systems to address the proposed aims. These studies should (i) be instrumental for our understanding of the diverse roles of the COMPASS family in the regulation of the pattern of H3K4 methylation and how they regulate development and differentiation; and (ii) have a fundamental impact on our understanding of how mutations within the COMPASS family result in cancer. This information has the potential of proving helpful to investigators attempting to design rational approaches for the treatment of cancer.