title: Pheromone- and RSP5-dependent ubiquitination of the G protein beta subunit Ste4 in yeast.
abstract: Ste4 is the β subunit of a heterotrimeric G protein that mediates mating responses in Saccharomyces cerevisiae. Here we show that Ste4 undergoes ubiquitination in response to pheromone stimulation. Ubiquitination of Ste4 is dependent on the E3 ligase Rsp5. Disrupting the activity of Rsp5 abolishes ubiquitination of Ste4 in vivo, and recombinant Rsp5 is capable of ubiquitinating Ste4 in vitro. We find also that Lys-340 is a major ubiquitination site on Ste4, as pheromone-induced ubiquitination of the protein is prevented when this residue is mutated to an arginine. Functionally, ubiquitination does not appear to regulate the stability of Ste4, as blocking ubiquitination has no apparent effect on either the abundance or the half-life of the protein. However, when presented with a concentration gradient of pheromone, Ste4(K340R) mutant cells polarize significantly faster than wild-type cells, indicating that ubiquitination limits pheromone-directed polarized growth. Together, these findings reveal a novel stimulus-dependent posttranslational modification of a Gβ subunit, establish Ste4 as a new substrate of the E3 ligase Rsp5, and demonstrate a role for G protein ubiquitination in cell polarization.
authors.affiliation: Department of Biology, Saint Louis University, St. Louis, Missouri 63103, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Zhu
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Torres
authors.forename: Joshua B
authors.initials: JB
authors.lastname: Kelley
authors.forename: Henrik G
authors.initials: HG
authors.lastname: Dohlman
authors.forename: Yuqi
authors.initials: Y
authors.lastname: Wang
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D044387:GTP-Binding Protein beta Subunits / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010675:Pheromones / Q000502:physiology*; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology*
chemical_list: D017931:DNA Primers; D044387:GTP-Binding Protein beta Subunits; D010675:Pheromones; D029701:Saccharomyces cerevisiae Proteins; C067576:Ste4 protein, S cerevisiae