RESUMEN
In humans, a mutation in the tyrosyl-DNA phosphodiesterase (Tdp1) is responsible for the recessively inherited syndrome spinocerebellar ataxia with axonal neuropathy (SCAN1). Tdp1 is a well-conserved DNA repair enzyme, which processes modified 3' phospho-DNA adducts in vitro. Here, we report that in the yeast Schizosaccharomyces pombe, tdp1 mutant cells progressively accumulate DNA damage and rapidly lose viability in a physiological G0/quiescent state. Remarkably, this effect is independent of topoisomerase I function. Moreover, we provide evidence that Tdp1, with the polynucleotide kinase (Pnk1), processes the same naturally occurring 3'-ends, produced from oxidative DNA damage in G0. We also found that one half of the dead cells lose their nuclear DNA. Nuclear DNA degradation is genetically programmed and mainly depends on the two DNA damage checkpoint responses, ATM/Tel1 and ATR/Rad3, reminiscent to programmed cell death. Diminishing the respiration rate or treating cells with a low concentration of antioxidants rescues the quiescent tdp1 mutant cells. These findings suggest that mitochondrial respiration causes neuronal cell death in the SCAN1 syndrome and in other neurological disorders.
Asunto(s)
División Celular , Daño del ADN , Hidrolasas Diéster Fosfóricas/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/citología , Schizosaccharomyces/enzimología , Apoptosis , Núcleo Celular/enzimología , Roturas del ADN de Cadena Simple , Reparación del ADN , Viabilidad Microbiana , Mitocondrias/enzimología , Mutación/genética , Oxidación-Reducción , Fenotipo , Polinucleótido 5'-Hidroxil-Quinasa/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Fase de Descanso del Ciclo CelularRESUMEN
Programmed cell death is a term which refers to a genetic decision of self-killing or suicide of a cell. Programmed cell death is not restricted to multicellular organisms and was described in a wide range of unicellular eukaryotes, indicating phylogenetically conserved functions, that participate in an adaptive response to cellular stress. Here we review and discuss our observations recently published in the EMBO Journal,(1) that non-dividing fission yeast, Schizosaccharomyces pombe, exhibits a DNA damage response leading to cell death. We found that Tdp1 protects quiescent S. pombe cells against oxidative DNA damage. Tdp1 is a well-conserved tyrosyl-DNA phosphodiesterase required for single-strand break DNA repair, the mutation of Tdp1 is responsible for the recessively inherited syndrome spinocerebellar ataxia with axonal neuropathy (SCAN1) in humans. We found that tdp1 mutant yeast cells grow, as well as the wild-type cells, during the vegetative state, but progressively die in the quiescent state. We showed that, in the absence of Tdp1, the accumulation of unrepaired oxidative DNA damage triggers a genetic response, leading to checkpoint-dependent (ATM/ATR) nuclear DNA degradation, reminiscent of apoptosis. Our results indicate that the reactive oxygen species (ROS) produced during mitochondrial respiration are the main DNA damaging agents in the physiological quiescent state.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Muerte Celular/fisiología , Daño del ADN/fisiología , Proteínas de Unión al ADN/metabolismo , Hidrolasas Diéster Fosfóricas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/fisiología , Proteínas Supresoras de Tumor/metabolismo , Apoptosis , Proteínas de la Ataxia Telangiectasia Mutada , Proteínas de Ciclo Celular/genética , Quinasa de Punto de Control 2 , Roturas del ADN de Cadena Simple , Reparación del ADN/fisiología , Proteínas de Unión al ADN/genética , Oxidación-Reducción , Hidrolasas Diéster Fosfóricas/genética , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Especies Reactivas de Oxígeno/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Proteínas Supresoras de Tumor/genéticaRESUMEN
Protein kinases orthologous with Cak1 of Saccharomyces cerevisiae (ScCak1) appear specific to ascomycetes. ScCak1 phosphorylates Cdc28, the cyclin-dependent kinase (CDK) governing the cell cycle, as well as Kin28, Bur1 and Ctk1, CDKs required for the transcription process performed by RNA polymerase II (RNA Pol II). Using genetic methods, we found that Cak1 genetically interacts with Paf1 and Ctr9, two components belonging to the PAF1 elongation complex needed for histone modifications, and with Ssu72, a protein phosphatase that dephosphorylates serine-5 phosphate in the RNA Pol II C-terminal domain. We present evidence suggesting that the interactions linking Cak1 with the PAF1 complex and with Ssu72 are not direct but mediated via Ctk1 and Bur1. We discuss the possibility that Ssu72 intervenes at the capping checkpoint step of the transcription cycle.