An analysis of genetic regulation of phospholipid synthesis will be conducted in the unicellular eukaryote, Saccharomyces cerevisiae. A major goal is to develop a comprehensive understanding of the regulatory cascade that controls transcriptional regulation of phospholipid biosynthetic structural genes in response to precursor availability and ongoing phospholipid synthesis. The studies will involve isolation and characterization of mutants with defects in the synthesis and/or regulation of phospholipids. The proposed research also involves a detailed analysis of the transcriptional regulation of structural genes encoding phospholipid biosynthetic enzymes. Four structural genes and three regulatory genes have been isolated and will be used in this analysis. The interaction of the trans-acting regulatory gene products with cis-acting elements in the promoters of the structural genes will be characterized in detail. The cloned structural genes will also be used to facilitate studies of the assembly of the phospholipid biosynthetic enzymes into their specific locations in the cellular membranes. Little is known about the mechanisms by which eukaryotic cells control the synthesis of membrane lipids in coordination with ongoing membrane biogenesis. Yet phospholipids, particularly inositol-containing phospholipids, have been implicated in complex signalling processes which play a role in controlling cell growth and proliferation in higher eukaryotes. Yeast cells synthesize a typically eukaryotic mixture of phospholipids, using pathways which are similar to those in higher eukaryotes. The organism is genetically tractable, and can be manipulated using powerful molecular genetics. Therefore, S. cerevisiae, provides an unusual opportunity to develop a comprehensive understanding of the regulatory mechanisms controlling phospholipid synthesis during membrane biogenesis and cell growth.
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