INO80-dependent chromatin remodeling regulates early and late stages of mitotic homologous recombination.
ABSTRACT Chromatin remodeling is emerging as a critical regulator of DNA repair factor access to DNA damage, and optimum accessibility of these factors is a major determinant of DNA repair outcome. Hence, chromatin remodeling is likely to play a key role in genome stabilization and tumor suppression. We previously showed that nucleosome eviction near double-strand breaks (DSBs) in yeast is regulated by the INO80 nucleosome remodeling complex and is defective in mutants lacking the Arp8 subunit of INO80. In the absence of homologous donor sequences, RPA recruitment to a DSB appeared normal in arp8Delta, but Rad51 recruitment was defective. We now show that the early strand invasion step of homologous recombination (HR) is markedly delayed in an arp8Delta haploid, but there is only a minor defect in haploid HR efficiency (MAT switching). In an arp8Delta diploid, interhomolog DSB repair by HR shows a modest defect that is partially suppressed by overexpression of Rad51 or its mediator, Rad52. In wild type cells, DSB repair typically results in gene conversion, and most gene conversion tracts are continuous, reflecting efficient mismatch repair of heteroduplex DNA. In contrast, arp8Delta gene conversion tracts are longer and frequently discontinuous, indicating defects in late stages of HR. Interestingly, when a homologous donor sequence is present, Rad51 is recruited normally to a DSB in arp8Delta, but its transfer to the donor is delayed, and this correlates with defective displacement of donor nucleosomes. We propose that retained nucleosomes at donors destabilize heteroduplex DNA or impair mismatch recognition, reflected in delayed strand invasion and altered conversion tracts.
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ABSTRACT: The mystery of nuclear actin has puzzled biologists for decades largely due to the lack of defined experimental systems. However, the development of actin-containing chromatin-modifying complexes as a defined genetic and biochemical system in the past decade has provided an unprecedented opportunity to dissect the mechanism of actin in the nucleus. Although the established functions of actin mostly rely on its dynamic polymerization, the novel finding of the mechanism of action of actin in the INO80 chromatin-remodeling complex suggests a conceptually distinct mode of actin that functions as a monomer. In this review we highlight the new paradigm and discuss how actin interaction with chromatin suggests a fundamental divergence between conventional cytoplasmic actin and nuclear actin. Furthermore, we provide how this framework could be applied to investigations of nuclear actin in other actin-containing chromatin-modifying complexes.Trends in cell biology 11/2013; · 12.12 Impact Factor
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ABSTRACT: Homologous recombination (HR) contributes to maintaining genome integrity by facilitating error-free repair of DNA double-strand breaks (DSBs) primarily during the S and G2 phases of the mitotic cell cycle, while non-homologous end joining (NHEJ) is the preferred pathway for DSB repair in G1 phase. The decision to repair a DSB by NHEJ or HR is made primarily at the level of DSB end resection, which is inhibited by the Ku complex in G1 and promoted by the Sae2 and Mre11 nucleases in S/G2 . The cell cycle regulation of homologous recombination is accomplished both at the transcription level and at the protein level through post-translational modification, degradation and subcellular localization. Cyclin-dependent kinase Cdc28 plays an established key role in these events, while the role of transcriptional regulation and protein degradation are less well understood. Here the cell cycle regulatory mechanisms for mitotic homologous recombination in Saccharomyces cerevisiae are reviewed and evolutionarily conserved principles are highlighted. This article is protected by copyright. All rights reserved.FEMS microbiology reviews 02/2014; · 10.96 Impact Factor
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ABSTRACT: Homologous recombination is crucial for genome stability and for genetic exchange. Although our knowledge of the principle steps in recombination and its machinery is well advanced, homology search, the critical step of exploring the genome for homologous sequences to enable recombination, has remained mostly enigmatic. However, recent methodological advances have provided considerable new insights into this fundamental step in recombination that can be integrated into a mechanistic model. These advances emphasize the importance of genomic proximity and nuclear organization for homology search and the critical role of homology search mediators in this process. They also aid our understanding of how homology search might lead to unwanted and potentially disease-promoting recombination events.Nature Reviews Molecular Cell Biology 05/2014; · 37.16 Impact Factor