The identification of Pcl1-interacting proteins that genetically interact with Cla4 may indicate a link between G1 progression and mitotic exit.
In budding yeast, Cla4 and Ste20, two p21-activated kinases, contribute to numerous morphogenetic processes. Loss of Ste20 or Cla4 individually confers distinct phenotypes, implying that they regulate different processes. However, loss of both proteins is lethal, suggesting some functional overlap. To explore the role(s) of Cla4, we and others have sought mutations that are lethal in a cla4 Delta strain. These mutations define >60 genes. Recently, both Ste20 and Cla4 have been implicated in mitotic exit. Here, we identify a genetic interaction between PHO85, which encodes a cyclin-dependent kinase, and CLA4. We further show that the Pho85-coupled G(1) cyclins Pcl1 and Pcl2 contribute to this Pho85 role. We performed a two-hybrid screen with Pcl1. Three Pcl1-interacting proteins were identified: Ncp1, Hms1, and a novel ATPase dubbed Epa1. Each of these proteins interacts with Pcl1 in GST pull-down experiments and is specifically phosphorylated by Pcl1.Pho85 complexes. NCP1, HMS1, and EPA1 also genetically interact with CLA4. Like Cla4, the proteins Hms1, Ncp1, and Pho85 appear to affect mitotic exit, a conclusion that follows from the mislocalization of Cdc14, a key mitotic regulator, in strains lacking these proteins. We propose a model in which the G(1) Pcl1.Pho85 complex regulates mitotic exit machinery.
Keniry
Megan E
ME
Department of Biology and Institute of Molecular Biology, University of Oregon, Eugene, Oregon 97403-1229, USA.
Kemp
Hilary A
HA
Rivers
David M
DM
Sprague
George F
GF
D000595:Amino Acid Sequence; D018844:Cyclin-Dependent Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D016193:G1 Phase; D005804:Genes, Lethal; D008938:Mitosis; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D004718:Saccharomycetales / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; C095212:CLA4 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D018844:Cyclin-Dependent Kinases