Regulation of the yeast amphiphysin homologue Rvs167p by phosphorylation.
The yeast amphiphysin homologue Rvs167p plays a role in regulation of the actin cytoskeleton, endocytosis, and sporulation. Rvs167p is a phosphoprotein in vegetatively growing cells and shows increased phosphorylation upon treatment with mating pheromone. Previous work has shown that Rvs167p can be phosphorylated in vitro by the cyclin-dependent kinase Pho85p complexed with its cyclin Pcl2p. Using chymotryptic phosphopeptide mapping, we have identified the sites on which Rvs167p is phosphorylated in vitro by Pcl2p-Pho85p. We have shown that these same sites are phosphorylated in vivo during vegetative growth and that phosphorylation at two of these sites is Pcl-Pho85p dependent. In cells treated with mating pheromone, the MAP kinase Fus3p is needed for full phosphorylation of Rvs167p. Functional genomics and genetics experiments revealed that mutation of other actin cytoskeleton genes compromises growth of a strain in which phosphorylation of Rvs167p is blocked by mutation. Phosphorylation of Rvs167p inhibits its interaction in vitro with Las17p, an activator of the Arp2/3 complex, as well as with a novel protein, Ymr192p. Our results suggest that phosphorylation of Rvs167p by a cyclin-dependent kinase and by a MAP kinase is an important mechanism for regulating protein complexes involved in actin cytoskeleton function.
Friesen
Helena
H
Department of Molecular and Medical Genetics, University of Toronto, Toronto, Canada, M5S 1A8.
Murphy
Kelly
K
Breitkreutz
Ashton
A
Tyers
Mike
M
Andrews
Brenda
B
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D007304:Insect Viruses; D008840:Microfilament Proteins; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D011994:Recombinant Proteins; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D051316:Wiskott-Aldrich Syndrome Protein
C477632:ARP2 protein, S cerevisiae; C102245:ARP3 protein, S cerevisiae; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D016213:Cyclins; D003598:Cytoskeletal Proteins; C480361:LAS17 protein, S cerevisiae; D008840:Microfilament Proteins; C090559:PCL2 protein, S cerevisiae; C082282:RVS167 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D051316:Wiskott-Aldrich Syndrome Protein; D018844:Cyclin-Dependent Kinases; C058605:PHO85 protein, S cerevisiae; C075580:FUS3 protein, S cerevisiae; D020928:Mitogen-Activated Protein Kinases