Mechanisms Regulating the Activation of Early Meiosis-Specific Genes

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In response to starvation the budding yeast, Saccharomyces cerevisiae, stops growing and dividing and enters into a process referred to as sporulation that leads to the production of haploid spores. The spores are resistant to harsh environmental conditions allowing the cells to survive for long periods of starvation or drought. Upon refeeding the spores can germinate as haploid cells and then engage in a mating process. Thus sporulation is required both for survival in harsh conditions and for sexual reproduction. The sporulation process requires a set of genes distinct from those expressed when the cells are actively growing and dividing. These sporulation-specific genes can be divided into families based upon the timing of their expression. The early genes are necessary for the initiation of the sporulation process. These genes are turned off during growth by the proteins Ume6, Sin3 and Rpd3. Upon starvation the IME1 gene is induced and the Ime1 protein binds to Ume6 to turn on the other early genes. One of these is IME2. When Ime2 protein accumulates it fully turns on the early genes. The IME1 gene is activated in response to starvation but only in cells that have fully functional mitochondria and are capable of respiration. Defective mitochondria or inhibition of respiration block IME1 transcription and the entire sporulation process. The objectives of this proposal are to determine how respiration competence regulates expression of the IME1 gene, and how Ime2 acts to drive elevated levels of early gene transcription required for sporulation. The role of Ime2 will be investigated by determining whether Ime2 directly modifies the repressor complex by phosphorylation. This will involve changing sites of phosphorylation in the repressor complex and determining whether its function is altered. The function of the mutated repressor will be tested for both its ability to turn off and turn on genes that it regulates. Additionally, The ability of Ime2 to control the translation of early sporulation-specific proteins will also be investigated as a potential mechanism that controls early genes. This will involve investigating whether there are any early gene mRNAs that are translated only when Ime2 is present. The link between respiration and activation of the IME1 gene will be investigated by making changes to the IME1 promoter and determining what parts respond to respiration competence. The relationship will be further investigated through genetic screens to identify factors that induce IME1 in respiration competent cells or repress IME1 in respiration incompetent cells. Realization of the objectives of this proposal will provide a model for understanding transcriptional regulation during cellular differentiation and provide an analysis of transcription and translational regulation in cellular differentiation.