? Defects in chromatin remodeling factors and histone modifying enzymes are associated with tumorigenesis and other genetic disorders. Therefore it is critical to understand these activities and their influence on one another. The human chromatin remodeling complex RSF (remodeling and spacing factor) is composed of two subunits, the novel protein Rsf-1 and the ISWI-type ATPase hSNF2H. RSF does not require exogenous histone chaperones for nucleosome assembly and spacing. The influence of specific histone modifications on the function of the individual subunits has not been determined; this will be accomplished through the use of native chemical ligation to synthesize site-specifically modified histones, as well as histones incorporating biophysical probes. In particular, the influence of these modifications on nucleosome assembly versus spacing will be assessed. The timing of the coupling of ATP hydrolysis to chromatin assembly events will also be examined. The function of Rsf-1 will be probed through mutagenesis, initially of the PHD zinc finger domain common to many chromatin-associated proteins. Rsf-1 and its PHD domain will be probed for their ability to bind phosphoinositols, and if so, for their influence on RSF function. ? ?
