The overall goals of our program of research are to understand temporal and spatial regulation of gene expression during endospore formation in the Gram-positive bacterium Bacillus subtilis and to understand how the programmed expression of sporulation genes is kept in register with the morphological events accompanying sporogenesis. Central to an investigation of these issues are five developmental-specific, RNA polymerase sigma factors and several auxiliary transcription factors that govern the program of sporulation gene expression. We propose to investigate how sigma factors mediate promoter recognition, how the appearance of sigma and auxiliary transcription factors is regulated temporally and spatially, which sets of genes are controlled by each factor and what kinds of feed-back mechanisms coordinate the appearance or activity of the factors with the course of sporogenesis. Specifically, we will"""""""" identify contact sites between a recognition helix for promoter """"""""- 10"""""""" sequences in the sporulation sigma factor sigmaH and bases in cognate promoter, investigate the role of sigmaH in early sporulation gene expression, test the proposal that sigmaH governs the transcription of the gene for the sporulation sigma factor sigmaF, identify genes under the control of sigmaF and modify the promoter recognition specificity of sigmaF, study the roles of sigmaE, sigmaK and transcription factors SpoIIID, spoVP and GerE in cascade regulation of spore coat genes, investigate the mechanistic basis for a rearrangement in the mother-cell chromosome that generates the composite structural gene for the mother-cell sigma factor sigmaK, investigate the possible existence of a pro-protein precursor to sigmaK and test the hypothesis that the """"""""morphological"""""""" coupling by the isolation of bypass mutations that permit coat gene expression in mutants blocked in forespore gene expression, and, finally, use immunoelectron microscopy to localize specific developmental proteins in spores and in sporulating cells. These goals address basic issues of temporal, spatial and morphological control of gene expression that are common to developmental systems of many kinds, including complex systems of norma and abnormal development in higher organisms.
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