Regulation of yeast protein kinase C activity by interaction with the small GTPase Rho1p through its amino-terminal HR1 domain.
Protein kinase C from Saccharomyces cerevisiae (Pkc1p) constitutes a prototypic member of the protein kinase C superfamily, as it shares all the conserved regions scattered among the isoenzymes of higher eukaryotes. The functional significance of some of the conserved domains in the yeast enzyme has not yet been investigated. We examined strains carrying a partial deletion in the amino-terminal region of the enzyme, which is homologous to the HR1 of the protein kinase C-related kinases. This strain was sensitive to the presence of caffeine, Calcofluor white and Congo red, all drugs known to affect mutants defective in the signal transduction pathway ensuring cellular integrity in which Pkc1p is a central component. Isolation of a single point mutation in HR1A, which shares the sensitivity to the drugs mentioned, confirmed the importance of this region for proper regulation of protein kinase C activity in vivo. Two-hybrid analysis provided evidence for an interaction of the small GTPase Rho1p with the HR1A region, in addition to the reported interaction of this protein with the C1 region of Pkc1p. MAP kinase phosphorylation assays indicate that this Rho1p-Pkc1p/HR1A interaction does not result in an activation of the kinase cascade. The intragenic lethality of mutants affected in both HR1A and the C1 domain reported in this work implies an essential role for Rho1p-Pkc1p interaction in yeast.
Schmitz
Hans-Peter
HP
Institut für Mikrobiologie, Heinrich-Heine-Universität Düsseldorf, Universitätsstr. 1, Geb. 26.12, Germany.
Lorberg
Anja
A
Heinisch
Jürgen J
JJ
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001557:Benzenesulfonates / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology; D003224:Congo Red / Q000494:pharmacology; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010766:Phosphorylation; D017354:Point Mutation; D025941:Protein Interaction Mapping; D011493:Protein Kinase C / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021122:Protein Subunits; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000187:drug effects; D013329:Structure-Activity Relationship; D020741:rho GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*
D001557:Benzenesulfonates; D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins; D002110:Caffeine; D003224:Congo Red; C007061:C.I. Fluorescent Brightening Agent 28; D011493:Protein Kinase C; C051302:RHO1 protein, S cerevisiae; D020741:rho GTP-Binding Proteins