GENETIC/BIOCHEM STUDIES OF CHROMATIN REMODELING FACTORS

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Nucleosomal histones and other components of chromatin can act as potent inhibitors of transcription by blocking the accessibility of both regulatory proteins and the general transcription machinery to DNA. Recent genetic and biochemical studies have identified a family of ATPases that counteract the repressive effects of chromatin. These chromatin remodeling factors are the catalytic subunits of huge heterometric complexes (including the yeast SWI/SNF, Drosophila BRM and human BRG1/HBRM complexes) that play highly conserved roles in eukaryotic cells. We are interested in the role of chromatin remodeling factors in Drosophila development, where alterations in chromatin structure are critical for the control of cell fate. This proposal is focused on two highly related Drosophila genes, brahma (brm) and imitation-SWI(ISWI). brm and ISWI encode the catalytic subunits of distinct chromatin remodeling complexes: the BRM complex and the nucleosome-remodeling fctor (NURF). brm plays an important role in the control of cell fate, but the biological function of ISWI is unknown. The proposed research is designed to elucidate the roles of brm and ISWI in animal development and clarify their mechanism of action in vivo. To gain an understanding of the mechanism of action of the BRM, we will identify the proteins with which it interacts in vivo. This will be accomplished by identifying the subunits of the BRM complex and screening for enhancers and suppressors of a dominant-negative brm mutation. To determine if brm and ISWI play similar roles in development, we will characterize the phenotype of loss-of-function and dominant-negative ISWI mutations. It has been proposed that BRM and ISWI may facilitate the binding of regulatory proteins to DNA by altering local chromatin structure. We will test this model by examining whether mutations in brm or ISWI affect the association of Polycomb and trithorax group proteins with polytene chromosomes. Recent studies have suggested that human proteins related to BRM are involved in development, viral integration, cell cycle control and cancer. The ability to conduct genetic studies of chromatin remodeling factors in Drosophila thus represents an excellent opportunity to increase our understanding of their roles in transcription, development and disease.