Project Summary/Abstract PI/PD: Pryciak, Peter M. Proper cell function and behavior depends on the ability to respond to signals in the extracellular environment and make appropriate decisions about whether or not to proliferate. In eukaryotic cells, responses to external signals are commonly initiated at the plasma membrane and then disseminated throughout the cell by signal transduction pathways, which control both cytoplasmic and nuclear events including gene expression. While many signaling pathways and their molecular components have been identified, some well-studied systems offer unique opportunities to understand how the molecular and biochemical properties of the individual pathway components shape the overall response of the system, and in a manner that integrates both positive and negative regulatory effects. This proposal explores such issues by using the mating reaction of the yeast Saccharomyces cerevisiae as a model system to understand fundamental aspects of eukaryotic signal transduction, via a molecular genetic and cell biological approach. The response to yeast mating pheromones involves a dynamic assembly of plasma membrane-localized signaling complexes, which include proteins found ubiquitously from yeast to humans such as a heterotrimeric G protein, a MAP kinase cascade, and a scaffold protein. The long-term objective of this project is to gain a molecular understanding of how signaling through this pathway is initiated and propagated, with an emphasis on the role of subcellular localization, the balance of positive and negative regulation, and the control of transcriptional responses. One goal will be to investigate the organization of signaling proteins into discrete compartments at the plasma membrane, and how this impacts the efficiency and dynamics of signal transmission. Another project will probe the mechanisms that control the ability of a MAP kinase protein to activate a negative feedback loop by inhibiting the pathway scaffold protein. Also under investigation will be how the gene expression program activated by pheromone is controlled by antagonistic effects of of two different pathway MAP kinases. Overall, these studies will contribute to our general understanding of signal transduction, with relevance to the mechanisms by which both normal and diseased cells make decisions regarding differentiation or proliferation.