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Smc5/6 maintains stalled replication forks in a recombination-competent conformation.

ABSTRACT: The Smc5/6 structural maintenance of chromosomes complex is required for efficient homologous recombination (HR). Defects in Smc5/6 result in chromosome mis-segregation and fragmentation. By characterising two Schizosaccharomyces pombe smc6 mutants, we define two separate functions for Smc5/6 in HR. The first represents the previously described defect in processing recombination-dependent DNA intermediates when replication forks collapse, which leads to increased rDNA recombination. The second novel function defines Smc5/6 as a positive regulator of recombination in the rDNA and correlates mechanistically with a requirement to load RPA and Rad52 onto chromatin genome-wide when replication forks are stably stalled by nucleotide depletion. Rad52 is required for all HR repair, but Rad52 loading in response to replication fork stalling is unexpected and does not correlate with damage-induced foci. We propose that Smc5/6 is required to maintain stalled forks in a stable recombination-competent conformation primed for replication restart.

SUBMITTER: Irmisch A

PROVIDER: S-EPMC2634738 | biostudies-literature | 2009 Jan

REPOSITORIES: biostudies-literature

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Smc5/6 maintains stalled replication forks in a recombination-competent conformation.

Irmisch Anja A Ampatzidou Eleni E Mizuno Ken'ichi K O'Connell Matthew J MJ Murray Johanne M JM

The EMBO journal 20090101 2

The Smc5/6 structural maintenance of chromosomes complex is required for efficient homologous recombination (HR). Defects in Smc5/6 result in chromosome mis-segregation and fragmentation. By characterising two Schizosaccharomyces pombe smc6 mutants, we define two separate functions for Smc5/6 in HR. The first represents the previously described defect in processing recombination-dependent DNA intermediates when replication forks collapse, which leads to increased rDNA recombination. The second n ...[more]

PMID: 19158664

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Project description:Expansive growth of neural progenitor cells (NPCs) is a prerequisite to the temporal waves of neuronal differentiation that generate the six-layered neocortex, while also placing a heavy burden on proteins that regulate chromatin packaging and genome integrity. This problem is further reflected by the growing number of developmental disorders caused by mutations in chromatin regulators. ATRX gene mutations cause a severe intellectual disability disorder (α-thalassemia mental retardation X-linked (ATRX) syndrome; OMIM no. 301040), characterized by microcephaly, urogenital abnormalities and α-thalassemia. Although the ATRX protein is required for the maintenance of repetitive DNA within heterochromatin, how this translates to disease pathogenesis remain poorly understood and was a focus of this study. We demonstrate that Atrx(FoxG1Cre) forebrain-specific conditional knockout mice display poly(ADP-ribose) polymerase-1 (Parp-1) hyperactivation during neurogenesis and generate fewer late-born Cux1- and Brn2-positive neurons that accounts for the reduced cortical size. Moreover, DNA damage, induced Parp-1 and Atm activation is elevated in progenitor cells and contributes to their increased level of cell death. ATRX-null HeLa cells are similarly sensitive to hydroxyurea-induced replication stress, accumulate DNA damage and proliferate poorly. Impaired BRCA1-RAD51 colocalization and PARP-1 hyperactivation indicated that stalled replication forks are not efficiently protected. DNA fiber assays confirmed that MRE11 degradation of stalled replication forks was rampant in the absence of ATRX or DAXX. Indeed, fork degradation in ATRX-null cells could be attenuated by treatment with the MRE11 inhibitor mirin, or exacerbated by inhibiting PARP-1 activity. Taken together, these results suggest that ATRX is required to limit replication stress during cellular proliferation, whereas upregulation of PARP-1 activity functions as a compensatory mechanism to protect stalled forks, limiting genomic damage, and facilitating late-born neuron production.

Dataset Information

Smc5/6 maintains stalled replication forks in a recombination-competent conformation.

Publications

Smc5/6 maintains stalled replication forks in a recombination-competent conformation.

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