PAF1复合物介导白血病原癌蛋白——MLL融合蛋白定位的分子基础研究

项目名称： PAF1复合物介导白血病原癌蛋白——MLL融合蛋白定位的分子基础研究

项目编号： No.31500613

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 生物科学

项目作者： 刘艳丽

作者单位： 华中师范大学

项目金额： 20万元

中文摘要： MLL是一种组蛋白甲基转移酶，参与催化组蛋白H3K4甲基化，从而激活下游基因的表达。该基因常与多种易位基因形成重组基因，表达的MLL融合蛋白是驱动白血病发展的重要原癌蛋白。研究表明，MLL融合蛋白可通过与PAF1复合物（PAF1c）相互作用而被定位于调控基因，然而该相互作用的分子基础尚不清楚。本项目在前期工作的基础上，对PAF1c如何介导MLL融合蛋白定位的分子基础展开研究：利用Pull-down和ITC等方法定位MLL与PAF1c组分的结合位点；通过解析MLL与PAF1c组分复合物晶体结构，揭示MLL与PAF1c相互作用的结构特点；依据结构设计突变体，通过体内外实验验证突变体对MLL融合蛋白的生物学功能的影响。本项目研究结果将展示MLL融合蛋白-PAF1c互作的分子机制，为阻断MLL-PAF1互作的小分子药物的开发提供理论基础，进而为治疗由MLL融合蛋白所致的白血病奠定基础。

中文关键词： MLL融合蛋白；PAF1复合物；蛋白质结构；白血病；相互作用

英文摘要： MLL (mixed lineage leukemia) is a histone methyltransferase which methylates histone H3 lysine 4 in MLL complex. Rearrangements of MLL gene are one of the most common genetic alterations in human leukemia. The most common MLL rearrangements result in oncogenic fusion protein of N terminal 1400 residues of MLL in frame with a variety of different translocation partners. MLL fusion proteins are among the most potent oncoproteins driving leukemia development. Recent studies reported that MLL and its fusion proteins could be recruited to chromatin through a direct interaction between an N-terminal fragment of MLL and the PAF1 complex (RNA Polymerase II Associated Factor 1 complex, PAF1c), and disruption of this interaction eliminated MLL-AF9 (an MLL fusion protein) mediated immortalization and leukemogenesis. Therefore, this MLL-PAF1c interaction provides an attractive venue to screen therapeutic compounds for treating MLL associated leukemia. However, the molecular mechanism of this interaction is still elusive. In this study, we will pursue the structural and functional study of this interaction. We will conduct the protein-protein interaction mapping between MLL and components of PAF1c by Pull-down and ITC (Isothermal titration calorimetry). In fact, we have finished the interaction mapping between MLL and PAF1, one component of PAF1c, and found that the fragment of PAF1 covering residues 164-380 interacts with MLL (residues 1115-1201). To elucidate the molecular basis of MLL-PAF1c interaction, we would like to solve the crystal structure of binary complexes between MLL and associated components of PAF1c. Based on the solved crystal structures, we will carry out site-directed mutagenesis study to identify critical residues affecting the interaction of the MLL N-terminal fragment and the PAF1c components. Once we have identified functionally important residues, we will validate the biological relevance of these critical residues in MLL complex recruitment by cell based experiments. Overall, our study will shed light on the molecular mechanisms of MLL-PAF1c interaction and greatly aid in drug discovery for leukemia.

英文关键词： MLL fusion protein;PAF1 complex;protein structure;leukemia;interaction