Molecular Mechanisms for the Assembly and Regulation of the MLL1 Core Complex

描述（申请人提供）：混合谱系白血病蛋白-1 (MLL1) 催化组蛋白 H3 赖氨酸 4 (H3K4) 甲基化，这是调节 HOX 基因在造血和发育过程中所必需的表观遗传标记。破坏 MLL1 基因的易位存在于一组独特的急性白血病中，通常预示着不良预后。其他 MLL1 重排和扩增会增加 MLL1 的酶活性并且具有致癌性。 MLL1 包含一个进化上保守的约 130 个氨基酸的 SET 结构域，可催化 H3K4 甲基化。最近的研究表明 MLL1 的酶活性受到保守的蛋白质复合物（包括 WDR5、RbBP5 和 ASH2L）的调节。这些蛋白质形成独立的复合物，与 MLL1 结合并调节其单甲基化、二甲基化或三甲基化 H3K4 底物的能力，这种现象称为“产物特异性”。由于 H3K4 的不同甲基化水平与不同的转录结果相关，因此有必要了解 MLL1 产物特异性调节的分子机制。尽管 MLL1 具有重要的生物学作用及其与人类白血病的关系，但目前有关 MLL1 核心复合物酶活性的蛋白质结构特征的信息很少。这项研究的长期目标是全面表征 MLL1 核心复合物调节 H3K4 甲基化的机制。 该提案采用结构-功能方法来研究 MLL1 核心复合物酶活性的分子机制。我们获得了新颖的初步结果，表明 MLL1 SET 结构域的内在产物特异性是缓慢的单甲基转移酶，但在与 WDR5、RbBP5 和 Ash2L 复合时变成快速的二甲基转移酶。在这里，我们建议确定 MLL1 核心复合物 H3K4 二甲基化活性的机制。此外，我们将确定导致其组装的蛋白质结构特征。为了实现这些目标，我们结合分子、生物化学和生物物理方法来研究 MLL1 核心复合物对 H3K4 甲基化的调节。这些信息将增加我们对关键酶复合物以及它如何在控制真核生物转录激活的途径中进行调节的理解。这项研究很重要，因为它可能会带来更好的诊断和合理设计抑制 MLL1 酶活性的抗癌药物。

DESCRIPTION (provided by applicant): The Mixed Lineage Leukemia protein-1 (MLL1) catalyzes histone H3 lysine 4 (H3K4) methylation, which is an epigenetic mark essential for the regulation of HOX genes in hematopoiesis and development. Translocations that disrupt the MLL1 gene are present in a unique group of acute leukemias, often predicting a poor prognosis. Other MLL1 rearrangements and amplifications increase MLL1's enzymatic activity and are oncogenic. MLL1 contains an evolutionarily conserved ~130 amino acid SET domain that catalyzes H3K4 methylation. Recent studies indicate that the enzymatic activity of MLL1 is regulated by a conserved complex of proteins including WDR5, RbBP5, and ASH2L. These proteins form an independent complex that binds to MLL1 and regulates its ability to mono-, di-, or trimethylate H3K4 substrates, a phenomenon known as 'Product Specificity'. Since different levels of methylation of H3K4 are associated with different transcriptional outcomes, it is imperative to understand the molecular mechanisms by which the product specificity of MLL1 is regulated. Despite the important biological role of MLL1 and its involvement in human leukemia, there is currently little information about the protein-structural features that are responsible for the enzymatic activity of the MLL1 core complex. The long-term goal of this research is to fully characterize the mechanisms for the regulation of H3K4 methylation by the MLL1 core complex. This proposal takes a structure-function approach to investigate the molecular mechanisms for the enzymatic activity of the MLL1 core complex. We have obtained novel preliminary results indicating that the intrinsic product specificity of the MLL1 SET domain is that of a slow monomethyltransferase, but becomes a fast dimethyltransferase when in complex with WDR5, RbBP5, and Ash2L. Here we propose to determine the mechanisms responsible for H3K4 dimethylation activity of the MLL1 core complex. In addition, we will determine the protein-structural features that account for its assembly. To address these aims, we combine molecular, biochemical, and biophysical approaches to investigate the regulation of H3K4 methylation by the MLL1 core complex. This information will increase our understanding of a key enzyme complex and how it is regulated in the pathways that control transcriptional activation in eukaryotes. This investigation is important because it may lead to better diagnostics and the rational design of anti-cancer drugs that inhibit MLL1's enzymatic activity.