Transcriptional regulation plays an important role in cellular homeostasis, regulating internal rhythms of the cell and its response to the external milieu. The cellular response to nutrient deprivation requires coordination of gene expression to insure that appropriate enzymes and cofactors are synthesized to meet the metabolic needs of the organism. Diabetes and metabolic syndrome are examples of two human diseases that arise when the metabolic response to caloric intake is not appropriately regulated at multiple levels, include transcriptional ones. Although not a disease strictly speaking, aging is another condition whose age of onset and severity is affected by nutrient stress, or caloric restriction, acting in part through transcriptional pathways affecting chromatin organization. The stress imposed by nutrient deprivation has been particularly well-studied in the budding yeast Saccharomyces cerevisiae. As in multicellular eukaryotes, metabolic stress in yeast is sensed and transmitted through an enzyme cascade that includes a complex protein kinase, the AMP-activated protein kinase (AMPK). The activity of the kinase cascade is modulated by a protein phosphatase complex. The targets of the kinase and phosphatase complexes include transcription factors and chromatin components as well as enzymes catalyzing key metabolic reactions in carbon source utilization. One important nutrient stress is glucose deprivation. In yeast as in other organisms, glucose starvation requires the organism to utilize other less readily available sources of carbon. These alternative energy sources are metabolized by pathways that are not utilized when glucose is available, a process called carbon catabolite repression or simply glucose repression. AMPK is activated when glucose is not available, and its activity serves as a switch to turn on numerous genes whose activity is necessary to process poorer carbon substrates such as ethanol, glycerol, and lactate, or to breakdown reserve carbohydrates and lipids, or to utilize alternative fermentable carbon sources. In our previous work we characterized two important transcription factors, Adr1 and Cat8, which are activated by AMPK. In the present proposal we focus on the characterization of an apparently novel protein phosphatase complex involved in gene repression, the chromatin changes that influence Adr1 binding, and the identification and characterization of upstream pathways that regulate the binding and activity of Adr1. ? ?
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