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

点击翻译按钮获取中文摘要

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.