RESUMO
Pmk1, a fission yeast homologue of mammalian ERK MAPK, regulates cell wall integrity, cytokinesis, RNA granule formation and ion homeostasis. Our screen for vic (viable in the presence of immunosuppressant and chloride ion) mutants identified regulators of the Pmk1 MAPK signaling, including Cpp1 and Rho2, based on the genetic interaction between calcineurin and Pmk1 MAPK. Here, we identified the vic2-1 mutants carrying a mis-sense mutation in the cwg2(+) gene encoding a beta subunit of geranylgeranyltransferase I (GGTase I), which participates in the post-translational C-terminal modification of several small GTPases, allowing their targeting to the membrane. Analysis of the vic2-1/cwg2-v2 mutant strain showed that the localization of Rho1, Rho4, Rho5 and Cdc42, both at the plasma and vacuolar membranes, was impaired in the vic2-1/cwg2-v2 mutant cells. In addition, Rho4 and Rho5 deletion cells exhibited the vic phenotype and cell wall integrity defects, shared phenotypes among the components of the Pmk1 MAPK pathway. Consistently, the phosphorylation of Pmk1 MAPK on heat shock was decreased in the cwg2-v2 mutants, and rho4- and rho5-null cells. Moreover, Rho4 and Rho5 associate with Pck1/Pck2. Possible roles of Cwg2, Rho4 and Rho5 in the Pmk1 signaling will be discussed.
Assuntos
Alquil e Aril Transferases/metabolismo , Parede Celular/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Alquil e Aril Transferases/química , Alquil e Aril Transferases/genética , Proteínas de Ligação ao GTP/genética , Sistema de Sinalização das MAP Quinases , Mutação , Fosforilação , Estrutura Terciária de Proteína , Schizosaccharomyces/citologia , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/genética , Proteínas rho de Ligação ao GTP/genéticaRESUMO
Rapamycin and its derivatives have now emerged as an attractive therapeutic strategy with both immunosuppressant and antitumor properties. In addition, rapamycin has been proposed as a calorie restriction mimetic to extend the life span of various organisms. The fission yeast Schizosaccharomyces pombe (S. pombe) serves as a valuable genetic model system to study the mechanism(s) of drug action as well as to determine genetic contexts associated with drug sensitivity or resistance. Here, we identified genes that when deleted modulate the rapamycin-sensitive strains in S. pombe. We carried out a chemical genomics screen for rapamycin-sensitive mutants using the genome-deletion library which covers 95.3% of all nonessential fission yeast genes and confirmed 59 genes to be rapamycin sensitive. Gene Ontology (GO) enrichment analysis showed that strains sensitive to rapamycin are highly enriched in processes regulating tRNA modification and mitochondria as well as other ontologies, including cellular metabolic process, chromatin organization, cell cycle, signaling, translation, transport and other cellular processes. Analysis also showed that components of the Elongator complex are overrepresented in the sensitive strains. Here, the data obtained will provide valuable information for speculation on the actions of rapamycin as well as on TORC signaling, thereby presenting a strategy to enhance sensitivity to rapamycin.