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

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，这是在造血和发育过程中调控HOX基因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，通常预示着预后不良。其他MLL1的重排和扩增增加了MLL1的酶活性，并具有致癌作用。MLL1包含一个进化上保守的约130个氨基酸的SET结构域，催化H3K4甲基化。最近的研究表明，MLL1的酶活性受包括WDR5、RbBP5和ASH2L在内的保守蛋白复合体的调节。这些蛋白质形成一个独立的复合体，与MLL1结合，并调节其对H3K4底物单、二或三甲基化的能力，这种现象被称为“产品特异性”。由于不同水平的H3K4甲基化与不同的转录结果相关，因此了解MLL1产物特异性调控的分子机制是必要的。尽管MLL1具有重要的生物学作用并参与人类白血病，但目前关于MLL1核心复合物酶活性的蛋白质结构特征的信息很少。本研究的长期目标是充分表征MLL1核心复合物调控H3K4甲基化的机制。本研究采用结构-功能方法研究MLL1核心复合物酶活性的分子机制。我们已经获得了新的初步结果，表明MLL1 SET结构域的内在产物特异性是一个缓慢的单甲基转移酶，但当与WDR5、RbBP5和Ash2L复合物时，它成为一个快速的二甲基转移酶。在这里，我们建议确定负责MLL1核心复合物的H3K4二甲基化活性的机制。此外，我们将确定解释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、生化和生物物理方法来研究MLL1核心复合物对H3K4甲基化的调控。这些信息将增加我们对关键酶复合物的理解，以及它如何在控制真核生物转录激活的途径中受到调节。这项研究具有重要意义，因为它可能导致更好的诊断和合理设计抑制MLL1酶活性的抗癌药物。

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.