Rapamycin and other natural products affect regulation of the growth and differentiation of Saccharomyces cerevisiae

摘要

The yeast Saccharomyces cerevisiae relies on many signal transduction pathways to sense its environment, most notably the availability of nutrients. The cell uses these signals to regulate gene expression, growth, and morphology to optimize its growth and survival in a changing environment. Many microorganisms produce small, cell-permeable chemicals that inhibit competing organisms. Often, these compounds specifically target signal transduction pathways, impairing the ability of the organism to sense its environment or regulate its growth and morphology. These natural products have proven useful in studying signal transduction processes.;The macrolide rapamycin specifically inhibits the TOR genes, phosphatidylinositol 4-kinase homologs that have been conserved from yeast to humans. TOR plays a central role in sensing and responding to available nutrients. In this dissertation, we present studies that utilize rapamycin in combination with genetics, biochemistry, molecular biology and genome array technology to probe the function of TOR in regulating the cells response to nitrogen. Rapamycin inhibits pseudohyphal differentiation in response to scarce nitrogen. This inhibition occurs at a concentration of the drug which does not effect vegetative growth rate but causes a dramatic reduction in translation. Rapamycin in rich medium induces the expression of genes to utilize less optimal nitrogen sources. We find that both the Tap42 protein and one of the phosphatases that it regulates, Sit4, are involved in TOR regulation of pseudohyphal differentiation. We propose a model whereby oscillations in TOR activity cause oscillations in Sit4 activity as nitrogen is encountered, then consumed, contributing to pseudohyphal differentiation.;We also report findings with fusel alcohols which induce a growth pattern similar to pseudohyphal differentiation. We find many genetic similarities, but also distinct differences from pseudohyphal growth in response to low nitrogen. We conclude that fusel alcohols stimulate starvation responses through pathways both shared and distinct from those that regulate pseudohyphal growth. We also present studies which reveal the phosphatidylinositol 4-kinase Stt4 to be the biologically relevant target of wortmannin, a steroid-like compound which inhibits many family members of the phosphatidylinositol 3-kinase superfamily of enzymes. Using small, natural products has proven useful in understanding the signal transduction pathways that regulate gene expression, cell growth, and differentiation.