title: Rho5p is involved in mediating the osmotic stress response in Saccharomyces cerevisiae, and its activity is regulated via Msi1p and Npr1p by phosphorylation and ubiquitination.
abstract: Small GTPases of the Rho family act as molecular switches, and modulation of the GTP-bound state of Rho proteins is a well-characterized means of regulating their signaling activity in vivo. In contrast, the regulation of Rho-type GTPases by posttranslational modifications is poorly understood. Here, we present evidence of the control of the Saccharomyces cerevisiae Rho-type GTPase Rho5p by phosphorylation and ubiquitination. Rho5p binds to Ste50p, and the expression of the activated RHO5(Q91H) allele in an Deltaste50 strain is lethal under conditions of osmotic stress. An overexpression screen identified RGD2 and MSI1 as being high-copy suppressors of the osmotic sensitivity of this lethality. Rgd2p had been identified as being a possible Rho5p GTPase-activating protein based on an in vitro assay; this result supports its function as a regulator of Rho5p activity in vivo. MSI1 was previously identified as being a suppressor of hyperactive Ras/cyclic AMP signaling, where it antagonizes Npr1p kinase activity and promotes ubiquitination. Here, we show that Msi1p also acts via Npr1p to suppress activated Rho5p signaling. Rho5p is ubiquitinated, and its expression is lethal in a strain that is compromised for proteasome activity. These data identify Rho5p as being a target of Msi1p/Npr1p regulation and describe a regulatory circuit involving phosphorylation and ubiquitination.
authors.affiliation: Department of Biochemistry, McIntyre Medical Building, McGill University, 3655 Promenade Sir William Osler, Montreal, Quebec H3G 1Y6, Canada. robert.annan@mail.mcgill.ca
authors.forename: Robert B
authors.initials: RB
authors.lastname: Annan
authors.forename: Cunle
authors.initials: C
authors.lastname: Wu
authors.forename: Daniel D
authors.initials: DD
authors.lastname: Waller
authors.forename: Malcolm
authors.initials: M
authors.lastname: Whiteway
authors.forename: David Y
authors.initials: DY
authors.lastname: Thomas
mesh_terms: D056504:Chromatin Assembly Factor-1; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D009997:Osmotic Pressure; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013489:Suppression, Genetic; D054875:Ubiquitination; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D056504:Chromatin Assembly Factor-1; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; C061736:MSI1 protein, S cerevisiae; C438904:Rgd2 protein, S cerevisiae; C079506:STE50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C066958:NPR1 protein, S cerevisiae; D011494:Protein Kinases; C474618:RHO5 protein, S cerevisiae; D020741:rho GTP-Binding Proteins