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

混合谱系白血病蛋白-1（MLL 1）催化组蛋白H3赖氨酸4（H3 K4）甲基化，这是造血和发育中HOX基因调控所必需的表观遗传标记。破坏MLL 1基因的易位存在于一组独特的急性白血病中，通常预测预后不良。其他MLL 1重排和扩增增加MLL 1的酶活性，并且是致癌的。MLL 1含有一个进化上保守的约130个氨基酸的SET结构域，其催化H3 K4甲基化。最近的研究表明，MLL 1的酶活性是由一个保守的蛋白质复合物，包括WDR 5，RbBP 5和ASH 2L调节。这些蛋白质形成一个独立的复合物，与MLL 1结合，并调节其单，二或三甲基化H3 K4底物的能力，这种现象称为“产品特异性”。由于H3 K4的不同甲基化水平与不同的转录结果相关，因此必须了解MLL 1产物特异性调节的分子机制。尽管MLL 1的重要生物学作用及其在人类白血病中的参与，但目前关于负责MLL 1核心复合物的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MLL 1核心复合物调节H3 K4甲基化的机制。该建议采用结构-功能方法来研究MLL 1核心复合物的酶活性的分子机制。我们已经获得了新的初步结果，表明MLL 1 SET结构域的内在产物特异性是缓慢的单甲基转移酶，但当与WDR 5，RbBP 5和Ash 2L复合时，成为快速的二甲基转移酶。在这里，我们建议确定负责MLL 1核心复合物的H3 K4二甲基化活性的机制。此外，我们将确定蛋白质的结构特征，占其组装。为了实现这些目标，我们结合联合收割机的分子，生物化学和生物物理的方法来研究的MLL 1核心复合物的H3 K4甲基化的调节。这些信息将增加我们对一个关键酶复合物的理解，以及它是如何在真核生物中控制转录激活的途径中受到调控的。这项研究很重要，因为它可能导致更好的诊断和抑制MLL 1酶活性的抗癌药物的合理设计。

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.