RESUMO
Non-homologous end joining (NHEJ) and homologous recombination (HR) are the two major pathways of DNA double-strand break (DSB) repair and both are highly conserved from yeast to mammals. Nej1 has a role in DNA end-tethering at a DSB, and the Mre11/Rad50/Xrs2 (MRX) complex is important for its recruitment to the break. Nej1 and Dna2-Sgs1 interact with the C-terminal end of Mre11, which also includes the region where Rad50 binds. By characterizing the functionality of Nej1 in two rad50 mutants, which alter the structural features of MRX, we demonstrate that Nej1 inhibits the binding of Dna2 to Mre11 and Sgs1. Nej1 interactions with Mre11 promote tethering and inhibit hyper-resection, and when these events are compromised, large deletions develop at a DSB. The work indicates that Nej1 provides a layer of regulation to repair pathway choice and is consistent with its role in NHEJ.
Assuntos
Quebras de DNA de Cadeia Dupla , DNA Helicases/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Endodesoxirribonucleases/metabolismo , Exodesoxirribonucleases/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , DNA Fúngico/metabolismo , Complexos Multiproteicos/metabolismo , RecQ Helicases/metabolismo , Saccharomyces cerevisiaeRESUMO
Is it possible to extract tethering forces applied on chromatin from the statistics of a single locus trajectories imaged in vivo? Chromatin fragments interact with many partners such as the nuclear membrane, other chromosomes or nuclear bodies, but the resulting forces cannot be directly measured in vivo. However, they impact chromatin dynamics and should be reflected in particular in the motion of a single locus. We present here a method based on polymer models and statistics of single trajectories to extract the force characteristics and in particular when they are generated by the gradient of a quadratic potential well. Using numerical simulations of a Rouse polymer and live cell imaging of the MAT-locus located on the yeast Saccharomyces cerevisiae chromosome III, we recover the amplitude and the distance between the observed and the interacting monomer. To conclude, the confined trajectories we observed in vivo reflect local interaction on chromatin.
