title: Cdc24 regulates nuclear shuttling and recruitment of the Ste5 scaffold to a heterotrimeric G protein in Saccharomyces cerevisiae.
abstract: The Saccharomyces cerevisiae guanine nucleotide exchange factor Cdc24 regulates polarized growth by binding to Cdc42, a Rho-type GTPase that has many effectors, including Ste20 kinase, which activates multiple MAPK cascades. Here, we show that Cdc24 promotes MAPK signaling during mating through interactions with Ste5, a scaffold that must shuttle through the nucleus and bind to the beta subunit (Ste4) of a G protein for Ste20 to activate the tethered MAPK cascade. Ste5 was basally recruited to growth sites of G1 phase cells independently of Ste4. Loss of Cdc24 inhibited nuclear import and blocked basal and pheromone-induced recruitment of Ste5. Ste5 was not basally recruited and the MAPK Fus3 was not basally activated in the presence of a Cdc24 mutant (G168D) that still activates Cdc42, suggesting that Cdc24 regulates Ste5 and the associated MAPK cascade through a function that is not dependent on its guanine nucleotide exchange factor activity. Consistent with this, Cdc24 bound Ste5 and coprecipitated with Ste4 independently of Far1 and Ste5. Loss of Cdc24 decreased Ste5-Ste4 complex formation, and loss of Ste4 stimulated Cdc24-Ste5 complex formation. Collectively, these findings suggest that Cdc24 mediates site-specific localization of Ste5 to a heterotrimeric G protein and may therefore ensure localized activation of the associated MAPK cascade.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Yunmei
authors.initials: Y
authors.lastname: Wang
authors.forename: Weidong
authors.initials: W
authors.lastname: Chen
authors.forename: David M
authors.initials: DM
authors.lastname: Simpson
authors.forename: Elaine A
authors.initials: EA
authors.lastname: Elion
mesh_terms: D000199:Actins / Q000737:chemistry; D021581:Active Transport, Cell Nucleus; D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry* / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000378:metabolism / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D003593:Cytoplasm / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000737:chemistry; D005838:Genotype; D005982:Glutathione Transferase / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism / Q000502:physiology*; D015151:Immunoblotting; D047468:Immunoprecipitation; D020935:MAP Kinase Signaling System; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D008957:Models, Genetic; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012097:Repressor Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D013696:Temperature; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013997:Time Factors
chemical_list: D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D019086:Bridged Bicyclo Compounds, Heterocyclic; C073594:CDC24 protein, S cerevisiae; D018797:Cell Cycle Proteins; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; C081353:FAR1 protein, S cerevisiae; D005656:Fungal Proteins; D020662:Guanine Nucleotide Exchange Factors; D012097:Repressor Proteins; C080321:STE5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013844:Thiazoles; D053778:Thiazolidines; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase; D019204:GTP-Binding Proteins; C037067:latrunculin A