Phosphorylation of Rad55 on Serines 2, 8, and 14 Is Required for Efficient Homologous Recombination in the Recovery of Stalled Replication Forks

Herzberg, Kristina; Bashkirov, Vladimir I.; Rolfsmeier, Michael; Haghnazari, Edwin; McDonald, W. Hayes; Anderson, Scott; Bashkirova, Elena V.; Yates, John R.; Heyer, Wolf Dietrich

doi:10.1128/mcb.01317-06

Cited by 83 publications

(85 citation statements)

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“…4 A relatively small number of proteins have been shown to become phosphorylated by the DDR; these include Ddc2, Mec1s binding partner, 39 the ssDNA binding protein RPA, 40 the Mcm4 and Mcm6 components of the replicative helicase, 41 the DNA primase, which synthesizes the RNA primer during DNA replication, 42 and the Rad55 protein, which participates in homologous recombination. 43 Here we add Elg1 to this list. Since the S112 serine phosphorylated is an SQ site, usually recognized and phosphorylated by PI3KKs, and deletion of MEC1 abolishes the modification (Fig.…”

Section: Discussionmentioning

confidence: 99%

Regulation of Elg1 activity by phosphorylation

et al. 2015

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These authors contributed equally to this work.Keywords: ATM/Tel1, ATR/Mec1, DNA damage response, DNA repair, DNA replication, telomeres ELG1 is a conserved gene with important roles in the maintenance of genome stability. Elg1's activity prevents gross chromosomal rearrangements, maintains proper telomere length regulation, helps repairing DNA damage created by a number of genotoxins and participates in sister chromatid cohesion. Elg1 is evolutionarily conserved, and its Fanconi Anemia-related mammalian ortholog (also known as ATAD5) is embryonic lethal when lost in mice and acts as a tumor suppressor in mice and humans. Elg1 encodes a protein that forms an RFC-like complex that unloads the replicative clamp, PCNA, from DNA, mainly in its SUMOylated form. We have identified 2 different regions in yeast Elg1 that undergo phosphorylation. Phosphorylation of one of them, S112, is dependent on the ATR yeast ortholog, Mec1, and probably is a direct target of this kinase. We show that phosphorylation of Elg1 is important for its role at telomeres. Mutants unable to undergo phosphorylation suppress the DNA damage sensitivity of Drad5 mutants, defective for an error-free post-replicational bypass pathway. This indicates a role of phosphorylation in the regulation of DNA repair. Our results open the way to investigate the mechanisms by which the activity of Elg1 is regulated during DNA replication and in response to DNA damage.

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Section: Discussionmentioning

confidence: 99%

Regulation of Elg1 activity by phosphorylation

et al. 2015

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“…Deletions of DNA damage checkpoint genes RAD9, RAD24 or RAD17 recover the mph1 srs2 defect, although less efficiently than HR inactivation (see Table 3). Recent findings suggested that the DNA damage checkpoint activates recombinational DNA repair for recovery after replication fork stalling (Herzberg et al, 2006). Therefore, viability of mph1 srs2 spores with a perturbed DNA damage checkpoint pathway could indicate that HR is less active in these mutants.…”

Section: Discussionmentioning

confidence: 99%

Genetic evidence for a role of Saccharomyces cerevisiae Mph1 in recombinational DNA repair under replicative stress

Panico

Ede

Schildmann

et al. 2009

Yeast

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In yeast as in human, DNA helicases play critical roles in assisting replication fork progression. The Saccharomyces cerevisiae MPH1 gene, homologue of human FANCM, has been involved in homologous recombination and DNA repair. We describe a synthetic growth defect of an mph1 deletion if combined with an srs2 deletion that can result -depending on the genetic background -in synthetic lethality. The lethality is suppressed by mutations in homologous recombination (rad51, rad52, rad55, rad57 ) and in the DNA damage checkpoint (rad9, rad24, rad17 ). Importantly, rad54 and mph1, epistatic for damage sensitivity, are subadditive for spontaneous mutator phenotype. Therefore, Mph1 could be placed at the Rad51-mediated strand invasion process, with a function distinct from Rad54. Moreover, siz1 mutation is viable with mph1 and additive for DNA damage sensitivity. mph1 srs2 double mutants, isolated in a background where they are viable, are synergistically sensitive to DNA damage. Moderate overexpression of SGS1 partially suppresses this sensitivity. Finally, we observe an epistatic relationship in terms of sensitivity to camptothecin of mms4 or mus81 to mph1. Overall, our results support a role of Mph1 in assisting replication progression. We propose two models for the resumption of DNA synthesis under replicative stress where Mph1 is placed at the sister chromatid interaction step.

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“…This phenotype is a consequence of a persistent activation of the DNA damage checkpoint, because this phenotype can be overridden by addition of caffeine (Mankouri and Hickson, 2006). Interestingly, mutation of RAD52 also impairs S-phase progression in the presence of MMS (Oakley et al, 2002), and substitution of phosphorylation sites targeted by checkpoint kinases in Rad55 causes defects in cell cycle resumption after MMS treatment (Herzberg et al, 2006). Taken together, these findings suggest that impaired S-phase progression in the presence of MMS may be a general property of HRR-defective mutants.…”

Section: Shu1 Functions In Rad51-and Rad54-dependent Homologous Recommentioning

confidence: 99%

Shu Proteins Promote the Formation of Homologous Recombination Intermediates That Are Processed by Sgs1-Rmi1-Top3

Mankouri

Ngo

Hickson

2007

MBoC

125

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CSM2, PSY3, SHU1, and SHU2 (collectively referred to as the SHU genes) were identified in Saccharomyces cerevisiae as four genes in the same epistasis group that suppress various sgs1 and top3 mutant phenotypes when mutated. Although the SHU genes have been implicated in homologous recombination repair (HRR), their precise role(s) within this pathway remains poorly understood. Here, we have identified a specific role for the Shu proteins in a Rad51/Rad54-dependent HRR pathway(s) to repair MMS-induced lesions during S-phase. We show that, although mutation of RAD51 or RAD54 prevented the formation of MMS-induced HRR intermediates (X-molecules) arising during replication in sgs1 cells, mutation of SHU genes attenuated the level of these structures. Similar findings were also observed in shu1 cells in which Rmi1 or Top3 function was impaired. We propose a model in which the Shu proteins act in HRR to promote the formation of HRR intermediates that are processed by the Sgs1-Rmi1-Top3 complex. INTRODUCTIONHomologous recombination repair (HRR) is a well-conserved cellular process for the repair of single-strand DNA (ssDNA) gaps and double-strand DNA breaks (DSBs) that can arise during DNA synthesis or as a result of replication fork stalling during S-phase. Although many of the key proteins involved in HRR have been identified (reviewed in Paques and Haber, 1999;Sung et al., 2003;West, 2003;Krogh and Symington, 2004), in some cases their precise function(s) remains to be identified. Moreover, the mechanisms for suppression of inappropriate HRR in S-phase are only poorly defined. It is likely that HRR must be carefully regulated and/or executed in dividing cells, as inappropriate or excessive HR can lead to genome rearrangements and cancer in mammals. This is exemplified by the cancer-predisposition disorder, Bloom's syndrome, which is caused by mutation in the human BLM gene (reviewed in German, 1993). Because the BLM protein, in conjunction with its associated proteins, hTOPOIII␣ and hRMI1 (Johnson et al., 2000;Wu et al., 2000;Yin et al., 2005), can catalyze dissolution of HRR intermediates in vitro Plank et al., 2006;Raynard et al., 2006;Wu et al., 2006), it is likely that unprocessed and/or aberrantly processed HRR intermediates at least partly contribute to the cellular defects in Bloom's syndrome. Indeed, Bloom's syndrome cells classically demonstrate elevated levels of sister chromatid exchanges, mitotic recombination, and genome instability. Mutation of the BLM, hTOPOIII␣, or hRMI1 homologues in Saccharomyces cerevisiae (SGS1, TOP3, or RMI1, respectively) similarly causes sensitivity to genotoxic agents, hyper-recombination, and synthetic lethality with mutations in other genes also implicated in HRR (e.g., MUS81 and SRS2; Gangloff et al., 1994;Watt et al., 1996;Gangloff et al., 2000;Mullen et al., 2001;Fabre et al., 2002;Chang et al., 2005;Mullen et al., 2005). Furthermore, unresolved HRR intermediates have been directly visualized by two-dimensional (2D) gel electrophoresis in cells lacking Sgs1 or in cells w...

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Phosphorylation of Rad55 on Serines 2, 8, and 14 Is Required for Efficient Homologous Recombination in the Recovery of Stalled Replication Forks

Cited by 83 publications

References 85 publications

Regulation of Elg1 activity by phosphorylation

Regulation of Elg1 activity by phosphorylation

Genetic evidence for a role of Saccharomyces cerevisiae Mph1 in recombinational DNA repair under replicative stress

Shu Proteins Promote the Formation of Homologous Recombination Intermediates That Are Processed by Sgs1-Rmi1-Top3

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