The homeotic genes of the Antennapedia and bithorax complexes encode spatially-restricted transcription factors that direct the choice between alternative pathways of development. The regulation of homeotic gene transcription is thus critical to the control of cell fate in Drosophila. Our long-term goal is to understand, at the molecular level, how the spatial patterns of homeotic gene transcription are established and maintained throughout Drosophila development. Genetic studies have identified many of the genes that control the transcription of homeotic genes during development, including Polycomb, a repressor of homeotic genes. Although recent studies have suggested that Polycomb represses homeotic gene transcription by influencing chromatin structure, its exact mechanism of action is unknown. To identify additional regulators of homeotic gene transcription, we have screened for dominant suppressors of Polycomb mutations. One of the genes identified in these screens, brahma (brm), encodes an activator of homeotic genes that is structurally related to the yeast transcriptional activator SNF2/SWI2. Genes encoding proteins that are highly related to brm have also been identified in mice and humans. In yeast, SNF2/SWI2 assists DNA-binding regulatory proteins to overcome the repressive effects of chromatin on transcription. Based on the similarities between brm and SNF2/SWI2, we have proposed that brm activates the transcription of homeotic genes by assisting DNA-binding regulatory proteins to overcome the repressive effects of Polycomb on chromatin structure. The experiments described in this proposal are designed to critically examine this hypothesis, and clarify the molecular mechanism of brm action. We will determine the role of brm during embryonic, larval and pupal stages of Drosophila development using germ- line and somatic clonal analysis. To determine whether brm is a functional homolog of SNF2/SWI2, we will test the ability of the brm protein to function in vivo in yeast. Biochemical studies will be conducted to identify and characterize brm-associated proteins, and the interactions between brm and its target genes will be examined by polytene chromosome immunostaining. We will also study the role of the brm protein in transcriptional activation using a relatively simple in vitro system.
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