Characterizaton of the chromatin remodelling activity of the transcription factor c-Myb

转录因子c-Myb在控制造血细胞的增殖、分化和凋亡中起关键作用，并与白血病和某些非造血性肿瘤的发生有关，如乳腺癌和结肠癌。虽然已知许多Myb调控基因，并已鉴定出几种与Myb相互作用和协同作用的蛋白质，但目前我们对Myb如何在天然的染色质嵌入状态下刺激其靶基因的表达还知之甚少。这样的图像将显著提高我们对Myb在造血细胞中的作用的理解，并将有助于开发Myb的小分子抑制剂。这个项目的总体目标是了解Myb和转录机制之间的分子相互作用，这些转录机制与Myb目标基因在其天然的显色状态下的转录调控有关。在我们的初步工作中，我们研究了Myb对鸡mim-1基因染色质结构的影响，并在Myb反式激活结构域中发现了一个高度保守的区域，该区域在Myb对染色质的重塑中起着关键作用。我们的工作表明，这个区域被称为染色质重塑结构域(CRD)，参与与Myb本身以及几个染色质重塑蛋白的蛋白质-蛋白质相互作用，包括蛋白精氨酸甲基转移酶PRMT4，组蛋白乙酰转移酶Tip60和Menin，一种适配蛋白混合系白血病(MLL)组蛋白甲基转移酶复合体。在这个项目中，我们计划详细探索这个结构域及其分子相互作用如何控制Myb作为转录因子的活性。具体地说，我们将表征CRD和Myb其他部分介导的分子内相互作用，利用蛋白质组学方法寻找CRD的新相互作用伙伴，并研究CRD与Menin、Tip60和PRMT4相互作用的功能相关性。总体而言，我们预计这项工作将极大地促进我们对Myb作为转录因子的功能的理解。我们还预计，从长远来看，CRD分子相互作用的表征将允许我们建立用于开发Myb功能治疗性抑制剂的检测系统。

The transcription factor c-Myb plays a key role in the control of proliferation, differentiation and apoptosis of hematopoietic cells and has been implicated in the development of leukemia and of certain non-hematopoietic tumors, such as breast and colon cancer. Although a number of Myb-regulated genes are known and several proteins interacting and cooperating with Myb have been identified, we have presently only a rather incomplete picture of how Myb stimulates the expression of its target genes in their native, chromatin-embedded state. Such a picture would significantly improve our understanding of the role of Myb in hematopoietic cells and would aid in the development of small molecule inhibitors of Myb. The overall goal of this project is to understand the molecular interactions between Myb and the transcriptional machinery that are relevant for the transcriptional regulation of Myb target genes in their native, chromatinized state. In our preliminary work we have studied the impact of Myb on the chromatin structure of the chicken mim-1 gene, a well-characterized Myb target gene, and identified a highly conserved region within the Myb transactivation domain, that plays a crucial role in chromatin remodelling by Myb. Our work shows that this region, referred to as chromatin remodelling domain (CRD), participates in protein-protein interactions with Myb itself as well as with several chromatin remodelling proteins, including the protein arginine methyl-transferase PRMT4, the histone acetyl-transferase TIP60 and Menin, an adaptor protein Mixed lineage leukemia (MLL) histone methyl-transferase complexes. In this project we plan to explore in detail how this domain and its molecular interactions control the activity of Myb as a transcription factor. Specifically, we will characterize intramolecular interactions mediated by the CRD and other parts of Myb, employ proteomic approaches to identify novel interaction partners of the CRD, and investigate the functional relevance of the interaction of the CRD with Menin, TIP60 and PRMT4. Overall, we expect that this work will significantly advance our understanding of the function of Myb as a transcription factor. We also expect that the characterization of the molecular interactions of the CRD will, on the longer run, permit us to establish assay systems for the development of therapeutic inhibitors of Myb function.