The mating type (MAI) genes of Saccharomyces cerevisiae are part of a complex network of genetic and physiological controls of cell type, cell lineage, and differentiation. The expression of the MAT genes themselves involves their activation by transposition from a silent copy, or donor, locus to the MAT locus, where either a or Alpha information is expressed. The work that we have proposed is designed to understand the mechanism by which MAT genes are transposed. The homothallic switching of MAT genes is a substitution/recombination event in which a copy of a of Alpha sequences at HML or HMR is used to replace the information at MAT. This gene conversion process is initiated by a site-specific endonuclease, but the molecular mechanism by which strands are transferred is not known. One major goal of this project is to define the molecular events occurring during this transposition process, to identify and characterize intermediates of the reaction and to define what DNA sequences at MAT and the donor loci are required for this process. In this regard, the project also seeks to determine in what ways the switching of MAT genes is similar to other mitotic recombination events. A second major goal of this project is to understand how the MAT locus interacts with the donor loci. Little is understood about the ways that distant sequences can be brought together efficiently to participate in a recombination event. When MATa cells switch, they preferentially interact with the donor HML, on the left arm of the same chromosome, while MATAlpha cells selectively pair with HMR. DNA sequences adjacent to MAT that affect this pairing will be characterized, as will mutations that affect the normal donor preference. The ability of cells to undergo meiosis depends both on the presence of MATa and MATAlpha alleles and on physiological signals. Experiments are also proposed to characterize a large class of mutations in 22 genes that affect normal G1 cell cycle arrest when cells are starved for amino acids or grown to stationary phase (and are thus unable to sporulate). Preliminary experiments have suggested that these mutations affect a network of genes that affect protein synthesis, amino acid biosynthesis and cell cycle arrest.
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