The Role of Polarity Proteins in regulating MAP Kinase Pathways in Saccharomyces cerevisiae
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During nutrient limiting conditions budding yeast undergo filamentous growth. One of the pathways that regulate filamentous growth is called the filamentous MAPK (fMAPK) pathway. In response to nutrient limitation, cell surface receptor-like proteins Msb2p, Sho1p, and Opy2p activate intracellular MAP Kinase cascades through the Rho GTPase Cdc42p. For my Ph.D. work, I focused on how the Rho GTPase Cdc42p recognizes and integrates environmental stimuli to regulate the filamentous growth pathway (fMAPK). Through my thesis work, I discovered the mechanism through which the spatial landmarks (bud-site-selection proteins) regulate the fMAPK pathway. Positional landmarks that direct the small monomeric GTPase Rsr1p to bud sites regulate the fMAPK pathway corresponding to their roles in bud-site selection through Cdc24p, the guanine nucleotide exchange factor (GEF) for the Cdc42p. In line with this conclusion, we find both extrinsic and intrinsic compromise of bud-site selection also impacted fMAPK activity. Therefore, a surveillance mechanism monitors the spatial position of the incipient bud site in response to extrinsic and intrinsic stress and modulates the response through the fMAPK pathway. For the second part of my thesis, I examined how Rho GTPase Cdc42p and the p21- activated kinase (PAK) Ste20p regulate MAPK pathways to detect and respond to different stimuli. To explore how Cdc42p is directed to the pathways in which it operates, I designed a system to compare the Cdc42p-dependent MAPK pathways, the fMAPK, HOG and mating pathways in the same cell. This approach identified Cdc42pE100A was defective for interaction with Bem4p, a pathway-specific adaptor for the MAPK pathway that controls filamentous growth (fMAPK). Corresponding residues in Bem4p were identified that were defective for interaction with Cdc42p. The polarity scaffold Bem1p also regulated the fMAPK pathway. Bem4p, Bem1p, and bud-site-selection GTPase Rsr1p, functioned in an ordered sequence (Rsr1p -> Bem4p -> Bem1p). Collectively, the study provides a mechanism for how a Rho GTPase is activated in a particular pathway.