RESUMEN
Rho GTPases are conserved molecules that control cytoskeletal dynamics. These functions are expedited by Rho GEFs that stimulate the release of GDP to enable GTP binding, thereby allowing Rho proteins to initiate intracellular signaling. How Rho GEFs and Rho GTPases protect cells from DNA damage is unknown. Here, we explore the extreme sensitivity of a deletion mutation in the Rho1p exchange factor Rgf1p to the DNA break/inducing antibiotic phleomycin (Phl). The Rgf1p mutant cells are defective in reentry into the cell cycle following the induction of severe DNA damage. This phenotype correlates with the inability of rgf1Δ cells to efficiently repair fragmented chromosomes after Phl treatment. Consistent with this observation Rad11p (ssDNA binding protein, RPA), Rad52p, Rad54p and Rad51p, which facilitate strand invasion in the process of homology-directed repair (HDR), are permanently stacked in Phl-induced foci in rgf1Δ cells. These phenotypes are phenocopied by genetic inhibition of Rho1p. Our data provide evidence that Rgf1p/Rho1p activity positively controls a repair function that confers resistance against the anti-cancer drug Phl.
Asunto(s)
Roturas del ADN de Doble Cadena , Reparación del ADN , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Cromosomas Fúngicos/genética , Roturas del ADN de Doble Cadena/efectos de los fármacos , Reparación del ADN/efectos de los fármacos , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de los fármacos , Proteínas Fluorescentes Verdes/metabolismo , Recombinación Homóloga/efectos de los fármacos , Viabilidad Microbiana/efectos de los fármacos , Mutación/genética , Fleomicinas/farmacología , Schizosaccharomyces/efectos de los fármacos , Transducción de Señal/efectos de los fármacosRESUMEN
In fission yeast, the septation initiation network (SIN) ensures temporal coordination between actomyosin ring (CAR) constriction with membrane ingression and septum synthesis. However, questions remain about CAR regulation under stress conditions. We show that Rgf1p (Rho1p GEF), participates in a delay of cytokinesis under cell wall stress (blankophor, BP). BP did not interfere with CAR assembly or the rate of CAR constriction, but did delay the onset of constriction in the wild type cells but not in the rgf1Δ cells. This delay was also abolished in the absence of Pmk1p, the MAPK of the cell integrity pathway (CIP), leading to premature abscission and a multi-septated phenotype. Moreover, cytokinesis delay correlates with maintained SIN signaling and depends on the SIN to be achieved. Thus, we propose that the CIP participates in a checkpoint, capable of triggering a CAR constriction delay through the SIN pathway to ensure that cytokinesis terminates successfully.