SATB1-protein Interaction/function at the BcL2 mbr/MAR

描述（由申请人提供）：该奖项将允许候选人在西奥多G. Krontiris在City of Hope国家医疗中心。经过这段时间的指导培训，候选人将留在希望之城的研究科学家的位置，主要负责研究。这种培训和知识的获取是必不可少的，她的未来目标，旨在了解的Bcl 2 T（14;18）易位的机制，这是一个煽动因素，在滤泡性淋巴瘤的发病。Bcl 2主要断裂点区域（mbr）的紧下游是AT富集区域，其结合特异性AT富集序列结合蛋白1（SATB 1），一种“胸腺细胞特异性”和MAR相关蛋白。SATB 1过表达下调Bcl 2表达。在体内DNA酶I足迹连接介导的PCR（LM-PCR）揭示了组成型和细胞类型特异性的蛋白结合在Bcl 2的mbr，也改变了DNA螺旋构型依赖于SATB 1。最近，酵母双杂交系统（y2 h）被用于鉴定与SATB 1相互作用的许多蛋白质。这些蛋白质中的许多蛋白质的所声称的功能对于MAR相关蛋白质来说似乎是合乎逻辑的（即，染色质重塑和转录）。出乎意料的是，SATB 1表现出与SUMO-1（一种蛋白质修饰因子）和其他SUMO-1酶分子的排他性相互作用。我们的研究主要包括以下几个方面：（1）通过生物化学、生物物理学和免疫学方法，研究SATB 1与SUMO-1修饰蛋白以及选择性转录因子和染色质重塑因子的相互作用。(2)使用Bcl 2 mbr/MAR的靶向诱变来确定哪些DNA序列有助于MAR功能。使用这些靶向细胞系来建立SATB 1和MARBc 12对特异性蛋白结合的贡献，以及对Bcl 2 mbr/MAR处的染色质构象和DNA拓扑结构的贡献。我们预计这些研究的成功完成将阐明导致疾病状态的细胞特异性蛋白和DNA功能的机制。

DESCRIPTION (provided by applicant): This award will allow the candidate to acquire an additional year of mentored training in techniques of cell biology and imaging technology, and targeted mutagenesis, in the laboratory of Theodore G. Krontiris at the City of Hope National Medical Center. After this period of mentored training, the candidate will remain at the City of Hope in a Research Scientist position, with responsibilities directed primarily towards research. This training and acquisition of knowledge is essential for her future goals directed towards understanding the mechanism of the Bcl2 t(14;18) translocation which is an instigating factor in the onset of follicular lymphoma. Immediately downstream of the Bcl2 major breakpoint region (mbr) is an AT-rich region that binds special AT-rich sequence binding protein l (SATB1), a 'thymocyte-specific', and MAR associating protein. SATB 1 over-expression down-regulates Bcl2 expression. In vivo DNase I footprinting by ligation-mediated PCR (LM-PCR) revealed constitutive and cell-type specific protein binding at the Bcl2 mbr and also an altered DNA helical configuration dependent upon SATB1. More recently, the yeast two-hybrid system (y2h) was used to identify numerous proteins that interact with SATB 1. The purported function(s) of many of these proteins appears logical for MAR-associating proteins (i.e., chromatin remodeling and transcription). Unexpectedly, SATB 1 demonstrated exclusive interaction with SUMO-1, a protein modifying factor, and with additional SUMO-1 enzymatic molecules. We propose to extend our studies as follows: (1) Characterize, through biochemical, biophysical, and immunological approaches, the interaction of SATB1 with SUMO-1 modifying proteins and with select transcription and chromatin remodeling factors. (2) Use targeted mutagenesis of the Bcl2 mbr/MAR to establish which DNA sequences contribute to MAR function. Use these targeted cell lines to establish the contribution of SATB 1 and MARBc12 to specific protein binding, and to chromatin conformation and DNA topology at the Bcl2 mbr/MAR. We anticipate that successful completion of these studies will elucidate mechanisms that contribute to cell-specific protein and DNA functions that lead to disease state.