EXCEED THE SPACE PROVIDED. This application is for renewal of a grant that focuses broadly on how sterol biosynthesis is regulated in Saccharomyces. Past work funded by this grant led to the discovery of protein prenylation and its importance in the function of Ras proteins, and to the discovery of a protein prenyl transferase, and the discovery of a proteolytic processing step shared by prenylated proteins. Additional progress included the discovery of several genes of the sterol biosyntheticpathway and multiple levels of regulation. The proposed experiments capitalize on this solid foundation and should lead to breakthroughs on several important issues: First, the PI and his team have discovered a pair of membrane bound transcription factors that mediate transcriptional control of sterol biosynthetic genes in response to fluctuations in sterol levels. These transcription factors are not homologs of mammalian SREBPs and thus represent convergent evolution of a new mechanism to control sterol synthesis. The key issue at hand is how these transcription factors are activated. A second focus builds on the exhaustive foundation laid in the last period on all 127 combinations of mutant genes encoding the oxysterol binding protein family. This work established that the seven family members provide at least one common essential function, which includes endocytosis, and unique gene- specific functions. The functions of each family member will be explored, especially in sterol trafficking and membrane dynamics. Previous work established the existence of a pair of prenyl-protein proteases with roles in processing virtually all prenylated proteins. Hypotheses regarding the specific protein substrates for both the yeast and human versions of these proteins will be tested. Finally, these hypothesis-driven studies will be complemented by a judicious combination of focused phenotypic surveys and technology development. These efforts will evaluate the generality of the lessons from earlier studies and begin an extension of these studies to the pathogenic fungus, Histoplasma capsulatum. The dominance of sterol targets in antifungal therapies form the basis for a comprehensive evaluation of the contribution of all Saccharomyces genes to sterol metabolism, and extension of this work to Histoplasma PERFORMANCE SITE ========================================Section End===========================================
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