ANALYSIS OF THE CHROMATIN-REMODELING COMPLEX RSC

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DESCRIPTION (provided by applicant): Our objectives are to understand how factors that remodel and modify chromosome structure contribute to gene regulation. Chromatin plays a major role in regulating trascription, primarily by restricting or promoting the access of transcription factors to genes. Chromatin structural changes are utilized to properly regulate many genes important for cell differentiation and proliferation, and misregulation of this process underlies many cancers. The formation of chromatin structures and their interconversion involves the concerted action of large complexes that modify and reposition/remodel the basic unit of chromatin structure, termed the nucleosome. Our studies focus on the essential yeast remodeling complex RSC, which is highly similar to mammalian SWl/SNF complexes. Thus, studies on RSC will have direct relevance to human SWI/SNF function. Our specific aims focus on understanding how nucleosomes are repositioned, how nucleosome modifications affect this process, and how modification and repositioning/remodeling are coordinated:1) Determine how DNA translocation is utilized for chromatin remodeling. DNA translocation has emerged as the unifying property of remodelers. We will use pre-steady state kinetics and special nucleosome substrates to understand how translocation is applied to remodel nucleosomes.2) Determine the role of RSC bromodomain- and BAH-containing modules in RSC targeting in vivo and in RSC remodeling in vitro. RSC complex contains essential bromodomains, and two essential BAH domains, which likely assist in RSC recognition of modified nucleosomes and/or coordination with modifiers. We will identify the modifications recognized, and determine their affect on RSC binding and remodeling.3) Define the role of actin and actin-related proteins in chromatin regulating complexes. Actin-related proteins and actin are present in many remodelers and modifiers. We will investigate their functions as hetermeric modules, in regulating RSC and NuA4 (modifier) function.