Chemical genetic analysis of the budding-yeast p21-activated kinase Cla4p.
The p21-activated kinases (PAKs) are effectors for the Rho-family GTPase Cdc42p. Here we define the in vivo function of the kinase activity of the budding yeast PAK Cla4p, using cla4 alleles that are specifically inhibited by a cell-permeable compound that does not inhibit the wild-type kinase. CLA4 kinase inhibition in cells lacking the partially redundant PAK Ste20p causes reversible SWE1-dependent cell-cycle arrest and gives rise to narrow, highly elongated buds in which both actin and septin are tightly polarized to bud tips. Inhibition of Cla4p does not prevent polarization of F-actin, and cytokinesis is blocked only in cells that have not formed a bud before inhibitor treatment; cell polarization and bud emergence are not affected by Cla4p inhibition. Although localization of septin to bud necks is restored in swe1Delta cells, cytokinesis remains defective. Inhibition of Cla4p activity in swe1Delta cells causes a delay of bud emergence after cell polarization, indicating that this checkpoint may mediate an adaptive response that is capable of promoting budding when Cla4p function is reduced. Our data indicate that CLA4 PAK activity is required at an early stage of budding, after actin polarization and coincident with formation of the septin ring, for early bud morphogenesis and assembly of a cytokinesis site.
Weiss
E L
EL
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3202, USA.
Bishop
A C
AC
Shokat
K M
KM
Drubin
D G
DG
D000483:Alleles; D000595:Amino Acid Sequence; D002451:Cell Compartmentation; D018797:Cell Cycle Proteins; D002455:Cell Division / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D005656:Fungal Proteins / Q000302:isolation & purification; D047908:Intracellular Signaling Peptides and Proteins; D020930:MAP Kinase Kinase Kinases; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D029701:Saccharomyces cerevisiae Proteins; D004718:Saccharomycetales / Q000166:cytology*
D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D005656:Fungal Proteins; D047908:Intracellular Signaling Peptides and Proteins; C410684:PCD1 protein, Pyrenopeziza brassicae; D011720:Pyrazoles; D011743:Pyrimidines; D029701:Saccharomyces cerevisiae Proteins; C099685:SWE1 protein, S cerevisiae; D011505:Protein-Tyrosine Kinases; C095212:CLA4 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D020930:MAP Kinase Kinase Kinases; C078754:STE20 protein, S cerevisiae