Rad3-dependent phosphorylation of the checkpoint clamp regulates repair-pathway choice

Kai, Mihoko; Furuya, Kanji; Paderi, Francesca; Carr, Antony; Wang, Teresa S.‐F.

doi:10.1038/ncb1600

Cited by 34 publications

(31 citation statements)

References 26 publications

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“…S3C). This suggests that Rad9-Thr 225 and Hus1-Thr 47 may not be true phosphorylated residues but are required for the clamp structure or other functions (39). Consistent with this, mutation of the two sites to the phosphomimic residue glutamic acid made the cells even more sensitive (supplemental Fig.…”

Section: Methodssupporting

confidence: 63%

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The Phosphorylation Network for Efficient Activation of the DNA Replication Checkpoint in Fission Yeast

Yue

Singh

Wang

et al. 2011

Journal of Biological Chemistry

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Section: Methodssupporting

confidence: 63%

“…47 in Hus1. Phosphorylation of Rad9 has been studied before for Chk1 activation and regulation of the choice of DNA repair pathways (22,25,39). All together, eight potential phosphorylations have been uncovered in the replication checkpoint pathway of S. pombe.…”

Section: Methodsmentioning

confidence: 99%

The Phosphorylation Network for Efficient Activation of the DNA Replication Checkpoint in Fission Yeast

Yue

Singh

Wang

et al. 2011

Journal of Biological Chemistry

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“…In budding yeast, the ScDsrs2 mutation suppresses the damage sensitivity of ScDrad18, presumably because Dsrs2 relieves the inhibition of the HR pathway and allows it to substitute for the PRR pathway; however, this is not observed in fission yeast (Kai et al 2007). We observed no genetic interactions between Dsrs2 and hsk1-1312 or dfp1- , and no changes in the damage sensitivity of the double mutants.…”

Section: Resultsmentioning

confidence: 40%

“…While activation of translesion synthesis may be coupled to a polymerase switching event at the fork, evidence suggests that it occurs behind the replication fork as well [reviewed in Branzei and Foiani (2007); Lambert et al (2007)]. Several studies suggest that checkpoint proteins may be intimately involved in the decision between recombination, template switching, and translesion synthesis (Paulovich et al 1998;Kai and Wang 2003;Liberi et al 2005;Kai et al 2007). An intriguing observation links DDK kinases specifically to translesion synthesis.…”

mentioning

confidence: 99%

Fission Yeast Hsk1 (Cdc7) Kinase Is Required After Replication Initiation for Induced Mutagenesis and Proper Response to DNA Alkylation Damage

Dolan

Schmidt

et al. 2010

Genetics

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Genome stability in fission yeast requires the conserved S-phase kinase Hsk1 (Cdc7) and its partner Dfp1 (Dbf4). In addition to their established function in the initiation of DNA replication, we show that these proteins are important in maintaining genome integrity later in S phase and G2. hsk1 cells suffer increased rates of mitotic recombination and require recombination proteins for survival. Both hsk1 and dfp1 mutants are acutely sensitive to alkylation damage yet defective in induced mutagenesis. Hsk1 and Dfp1 are associated with the chromatin even after S phase, and normal response to MMS damage correlates with the maintenance of intact Dfp1 on chromatin. A screen for MMS-sensitive mutants identified a novel truncation allele, rad35 (dfp1-(1-519)), as well as alleles of other damage-associated genes. Although Hsk1-Dfp1 functions with the Swi1-Swi3 fork protection complex, it also acts independently of the FPC to promote DNA repair. We conclude that Hsk1-Dfp1 kinase functions post-initiation to maintain replication fork stability, an activity potentially mediated by the C terminus of Dfp1.T HE Hsk1 protein kinase, the fission yeast ortholog of Saccharomyces cerevisiae Cdc7, is a conserved protein essential for the initiation of DNA replication (Masai et al. 1995;Brown and Kelly 1998;Snaith et al. 2000). Data from many systems suggest that the kinase functions at individual replication origins to activate the prereplication complex (preRC) through phosphorylation of the MCM helicase and other subunits (reviewed in Forsburg 2004). In fission yeast, Hsk1 kinase activity is limited to S phase by its regulatory subunit Dfp1, which is transcriptionally and posttranslationally regulated to restrict its peak of activity to S phase (Brown and Kelly 1999;Takeda et al. 1999). The requirement for Dfp1 (in S. cerevisiae, Dbf4) is similar to the dependence of CDK kinases on cyclin activity; thus, the Ccd7 kinase family has been dubbed DDK (Dbf4-dependent kinases) ( Johnston et al. 1999;Duncker and Brown 2003). Hsk1 is a target of the Cds1 checkpoint kinase and undergoes Cds1-dependent phosphorylation during hydroxyurea (HU) treatment in vivo and in vitro (Snaith et al. 2000). Interestingly, deletion of Dcds1 partly rescues hsk1-1312 temperature sensitivity, which suggests that Hsk1 is negatively regulated by the replication checkpoint. In turn, Cds1 is poorly activated in hsk1 mutants after HU treatment, indicating that there may be a feedback loop linking these two kinases (Snaith et al. 2000;Takeda et al. 2001). hsk1 mutants are sensitive to HU treatment, with a phenotype suggesting a specific defect in recovery (Snaith et al. 2000).DDK kinases have substrates outside of the replication initiation pathway. Functional dissection of Schizosaccaromyces pombe Dfp1 identifies separate regions that are required for checkpoint response (N-terminal domain; Takeda et al. 1999;Fung et al. 2002), for centromere cohesion and replication (MIR domain;Bailis et al. 2003;Hayashi et al. 2009) and for proper response to alkylat...

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“…For instance, Rad9 of the S. pombe 9-1-1 complex, which forms a PCNA-like heterotrimeric clamp, associates with Mms2, and a mutant form of Rad9 incapable of interaction promotes mutagenesis in a TLS-dependent manner [133]. In budding yeast the phosphorylation by protein kinase Mec1 induces the re-localization of Rev1 and Polξ to sites of DNA double-strand breaks independently of mono-Ub PCNA [134].…”

Section: Regulated Access Of Y-family Polymerases To the Damage Sitementioning

confidence: 99%

Eukaryotic DNA damage tolerance and translesion synthesis through covalent modifications of PCNA

Andersen¹,

Xiao³

2007

Cell Res

185

187

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npg In addition to well-defined DNA repair pathways, all living organisms have evolved mechanisms to avoid cell death caused by replication fork collapse at a site where replication is blocked due to disruptive covalent modifications of DNA. The term DNA damage tolerance (DDT) has been employed loosely to include a collection of mechanisms by which cells survive replication-blocking lesions with or without associated genomic instability. Recent genetic analyses indicate that DDT in eukaryotes, from yeast to human, consists of two parallel pathways with one being error-free and another highly mutagenic. Interestingly, in budding yeast, these two pathways are mediated by sequential modifications of the proliferating cell nuclear antigen (PCNA) by two ubiquitination complexes Rad6-Rad18 and Mms2-Ubc13-Rad5. Damage-induced monoubiquitination of PCNA by Rad6-Rad18 promotes translesion synthesis (TLS) with increased mutagenesis, while subsequent polyubiquitination of PCNA at the same K164 residue by Mms2-Ubc13-Rad5 promotes error-free lesion bypass. Data obtained from recent studies suggest that the above mechanisms are conserved in higher eukaryotes. In particular, mammals contain multiple specialized TLS polymerases. Defects in one of the TLS polymerases have been linked to genomic instability and cancer. DNA damage toleranceIn the presence of spontaneous or carcinogen-induced DNA damage, living cells have to maintain and complete DNA synthesis or risk replication fork collapse. Since the process of DNA licensing is to ensure the genome is duplicated once and only once during each cell cycle, stalled or collapsed replication forks may not be able to restart, which often results in double-strand breaks (DSBs) and causes compromised genome integrity or cell death. In addition to highly conserved DNA repair pathways, all living organisms have evolved schemes to ensure continuation of DNA synthesis in the presence of damage. These schemes were originally termed DNA postreplication repair (PRR) due to observations of transient shortened nascent DNA structures following S phase in response to DNA damage. In bacteria and unicellular yeast, these shortened DNA segments can be measured by an alkaline sedimentation assay [1] or directly observed in electron micrographs [2]. In wild-type cells, these truncated DNA segments were restored to full length following a short recovery time. One typical experiment [1] involved the restoration of the nascent strand following UV exposure in nucleotide excision repair (NER)-deficient cells and was originally assumed to represent a mechanism of DNA repair. However, further investigation revealed that, although the nascent fragments were re-annealed, the original UV-induced pyrimidine dimers, which were responsible for the generation of single-strand gaps, often persisted in the genome [3,4]. It was argued that the replication-blocking lesion was not necessarily corrected, but rather transiently bypassed and carried over to the next generation. Perhaps it is more beneficial for the organi...

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Rad3-dependent phosphorylation of the checkpoint clamp regulates repair-pathway choice

Cited by 34 publications

References 26 publications

The Phosphorylation Network for Efficient Activation of the DNA Replication Checkpoint in Fission Yeast

The Phosphorylation Network for Efficient Activation of the DNA Replication Checkpoint in Fission Yeast

Fission Yeast Hsk1 (Cdc7) Kinase Is Required After Replication Initiation for Induced Mutagenesis and Proper Response to DNA Alkylation Damage

Eukaryotic DNA damage tolerance and translesion synthesis through covalent modifications of PCNA

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