Regulation of signal transduction by G protein β subunits in Saccharomyces cerevisiae

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Alternate title
  • Regulation of signal transduction by G protein [beta] subunits in Saccharomyces cerevisiae
Last Modified
  • March 21, 2019
Creator
  • Zeller, Corinne Eileen
    • Affiliation: School of Medicine, Department of Biochemistry and Biophysics
Abstract
  • All cells must be able to respond to extracellular signals and environmental changes by initiating appropriate cellular responses. Guanine nucleotide binding proteins (G proteins) are key intermediates in cellular signaling and play a role in responding to a variety of extracellular stimuli. The yeast Saccharomyces cerevisiae contains two G protein mediated signaling pathways, one for glucose sensing and a second for pheromone responsiveness. Yeast have many signaling pathways that are initiated and regulated in response to glucose, but only one is G protein mediated. In this pathway the Gα subunit Gpa2 is coupled to the Gpr1 receptor; upon glucose stimulation Gpa2 activates production of the second messenger cAMP. Heterotrimeric G proteins contain, by definition, three subunits: α, β, and γ. However, the Gβγ heterodimer that functions with Gpa2 to mediated glucose signaling has never been identified. I have shown that Asc1, a multifunctional protein that contains the 7WD repeat domain structure of known Gβ subunits, functions as the Gβ subunit in the glucose signaling pathway. As with other known Gβ proteins, Asc1 interacts directly with the Gα in a guanine nucleotide-dependent manner, and inhibits Gα guanine nucleotide exchange activity. In addition, Asc1 interacts with the effector enzyme adenylyl cyclase (Cyr1), and functions to diminish the production of cAMP in response to glucose stimulation. Thus while Gpa2 promotes glucose signaling through elevated production of cAMP, Asc1 restricts this same process. The RGS protein in the yeast S.cerevisiae pheromone response pathway, Sst2, is a divergent member of the R7 family of RGS proteins. The R7 family of RGS proteins is distinct from other RGS protein families because it has a unique binding partner, Gβ5 (the type five isoform of Gβ subunits). The Gβ5-RGS heterodimer functions like many other RGS proteins as a GTPase accelerating protein towards Gα subunits, but the role that the Gβ5 plays in this process is unknown. Therefore, I sought to determine if Sst2 has a Gβ5-like binding partner and if so, to use the pheromone response pathway as a tool to elucidate the function of the Gβ5 subunit in regulating G protein signaling. I identified the protein Prp4 as a candidate Gβ5 because it interacts with both Sst2 and the Gα Gpa1. However, I have been unable to definitively determine if Prp4 is a Gβ5-like protein. The Gβ subunit in the S. cerevisiae pheromone response pathway becomes rapidly phosphorylated upon pheromone stimulation. However, the kinase that performs the phosphorylation, and the functional consequences of this event, are unknown. I have used genetic and biochemical methods to try to determine the identity of the kinase, the specific sites of phosphorylation, and the functional outcomes of phosphorylation. The MAP kinase in the pheromone response pathway, Fus3, has been implicated as the kinase that phosphorylates Ste4, however direct evidence for this has been lacking. I have now shown that Fus3 is not the kinase; my results do indicate that the Ste4 kinase is downstream of Fus3 signaling. When first identified, it was believed that the sole function of the Gβγ subunit was to regulate the Gα subunit, by targeting the Gα to the plasma membrane and inhibiting
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  • In Copyright
Advisor
  • Dohlman, Henrik
Degree granting institution
  • University of North Carolina at Chapel Hill
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  • Open access
Date uploaded
  • October 19, 2010

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