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During replication stress, non-SMC element 5 (NSE5) is required for Smc5/6 protein complex functionality at stalled forks.

ABSTRACT: The Smc5/6 complex belongs to the SMC (structural maintenance of chromosomes) family, which also includes cohesin and condensin. In Saccharomyces cerevisiae, the Smc5/6 complex contains six essential non-Smc elements, Nse1-6. Very little is known about how these additional elements contribute to complex function except for Nse2/Mms21, which is an E3 small ubiquitin-like modifier (SUMO) ligase important for Smc5 sumoylation. Characterization of two temperature-sensitive mutants, nse5-ts1 and nse5-ts2, demonstrated the importance of Nse5 within the Smc5/6 complex for its stability and functionality at forks during hydroxyurea-induced replication stress. Both NSE5 alleles showed a marked reduction in Smc5 sumoylation to levels lower than those observed with mms21-11, a mutant of Mms21 that is deficient in SUMO ligase activity. However, a phenotypic comparison of nse5-ts1 and nse5-ts2 revealed a separation of importance between Smc5 sumoylation and the function of the Smc5/6 complex during replication. Only cells carrying the nse5-ts1 allele exhibited defects such as dissociation of the replisome from stalled forks, formation of fork-associated homologous recombination intermediates, and hydroxyurea sensitivity that is additive with mms21-11. These defects are attributed to a failure in Smc5/6 localization to forks in nse5-ts1 cells. Overall, these data support the premise that Nse5 is important for vital interactions between components within the Smc5/6 complex, and for its functionality during replication stress.

SUBMITTER: Bustard DE

PROVIDER: S-EPMC3322872 | biostudies-other | 2012 Mar

REPOSITORIES: biostudies-other

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During replication stress, non-SMC element 5 (NSE5) is required for Smc5/6 protein complex functionality at stalled forks.

Bustard Denise E DE Menolfi Demis D Jeppsson Kristian K Ball Lindsay G LG Dewey Sidney Carter SC Shirahige Katsuhiko K Sjögren Camilla C Branzei Dana D Cobb Jennifer A JA

The Journal of biological chemistry 20120202 14

The Smc5/6 complex belongs to the SMC (structural maintenance of chromosomes) family, which also includes cohesin and condensin. In Saccharomyces cerevisiae, the Smc5/6 complex contains six essential non-Smc elements, Nse1-6. Very little is known about how these additional elements contribute to complex function except for Nse2/Mms21, which is an E3 small ubiquitin-like modifier (SUMO) ligase important for Smc5 sumoylation. Characterization of two temperature-sensitive mutants, nse5-ts1 and nse5 ...[more]

PMID: 22303010

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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

During replication stress, non-SMC element 5 (NSE5) is required for Smc5/6 protein complex functionality at stalled forks.

Publications

During replication stress, non-SMC element 5 (NSE5) is required for Smc5/6 protein complex functionality at stalled forks.

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