Regulation of the GATA-type transcription factor Gln3p by the target of rapamycin protein (TOR)

Carbon and nitrogen are two basic nutrient sources used by eukaryotic cells to produce energy and synthesize biomolecules important for cell growth and proliferation. In response to the quality of these nutrients, cells can regulate expression of genes involved in different metabolic pathways. A classic example is nitrogen catabolite repression (NCR) in yeast Saccharomyces cerevisiae, where cells regulate expression of genes involved in the utilization and transport of available nitrogen nutrients. One primary NCR transcription factor is the GATA-type transcription activator Gln3p. This protein was shown to be sequestered in the cytoplasm by the prion-precursor Ure2p when cells are grown in preferred nitogren. In addition, it was shown that the target of rapamycin (TOR) protein-nitrogen and Snflp-glucose signaling pathways regulate Gln3p.; Rapamycin is a potent immunosuppressant and candidate drug for organ and bone marrow transplantation, treatment of autoimmune diseases, anti-tumorigenesis and fungal infections. The target of rapamycin (TOR) protein is a 220 Kda ataxia telangiectasia mutated (ATM)-related protein kinase that is inhibited by rapamycin when rapamycin is complexed with its immunophilin receptor FKBP 12. TOR is a conserved serine/threonine protein kinase and key player in nutrient-mediated signal transduction to control cell growth and proliferation. We recently discovered that TOR interacts with Gln3p and Ure2p. We also found that nitrogen starvation or inhibition of TOR by rapamycin causes rapid dephosphorylation and nuclear accumulation of Gln3p in vivo as well as expression of a wide range of NCR genes. In addition, TOR appears to be responsible for Gln3p phosphorylation and may also regulate Gln3p dephosphorylation. Our data indicate that the ability of TOR to regulate transcription via Gln3p is a major signaling mechanism important for rapamycin-sensitive cell growth as well as nitrogen nutrient signaling. To understand TOR function and its relevance for human disease, my dissertation research focuses on using the yeast Saccharomyces cerevisiae as a model organism for studying TOR and Gln3p, with the intent of discovering the nuclear import mechanism for Gln3p, the relevant structural domains of Gln3p important for its function, and the role of TOR, Ure2p, phosphorylation, and nutrients in the regulation of Gln3p and, ultimately, TOR-regulated gene expression.