Rad9 plays an important role in DNA mismatch repair through physical interaction with MLH1

He, Wei; Zhao, Yun; Zhang, Chunbo; An, Lili; Hu, Zhishang; Liu, Yuheng; Han, Lu; Bi, Lijun; Xie, Zenghong; Xue, Peng; Yang, Fuquan; Hang, Haiying

doi:10.1093/nar/gkn686

Cited by 43 publications

(49 citation statements)

References 68 publications

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“…40 Multiple interactions have also been reported between 9-1-1 subunits and the DNA mismatch repair factors MutS homolog 2 (MSH2), MSH3, MSH6, and MutL homolog 1 (MLH1), 41,42 and mismatch recognition by the MSH proteins is stimulated by the presence of 9-1-1. 41 Furthermore, human RAD9A and MSH6 colocalize in nuclear foci following DNA methylating agent treatment, and RAD9A focus formation is dependent upon the presence of MSH6.…”

Section: The 9-1-1 Complex Functions In Atr Activation and Dna Repairmentioning

confidence: 99%

“…Such a possibility would be consistent with the fact that RAD9A foci normally persist on the sex chromosome cores during pachynema, colocalizing with RAD51 foci at these sites. 15 Although RAD9A can interact with MLH1, 42 it seems unlikely that the persistent DSBs in Rad9a and Hus1 CKOs result from failed crossing over, as approximately normal numbers of pachytene MLH1 foci and bivalent diakinesis chromosomes were observed in the absence of HUS1. We also cannot exclude the possibility that HUS1 is involved in partner choice during the homology search, and that perhaps some of the DSB sites indicated by MLH1 foci in the Hus1 CKO are resolved by inter-sister chromatid rather than interhomolog chromosome recombination events.…”

Section: Rad9a and Hus1 Are Essential For A Subset Of Meiotic Recombimentioning

confidence: 99%

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Clamping down on mammalian meiosis

et al. 2013

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T he RAD9A-RAD1-HUS1 (9-1-1) complex is a PCNA-like heterotrimeric clamp that binds damaged DNA to promote cell cycle checkpoint signaling and DNA repair. While various 9-1-1 functions in mammalian somatic cells have been established, mounting evidence from lower eukaryotes predicts critical roles in meiotic germ cells as well. This was investigated in 2 recent studies in which the 9-1-1 complex was disrupted specifically in the mouse male germline through conditional deletion of Rad9a or Hus1. Loss of these clamp subunits led to severely impaired fertility and meiotic defects, including faulty DNA double-strand break repair. While 9-1-1 is critical for ATR kinase activation in somatic cells, these studies did not reveal major defects in ATR checkpoint pathway signaling in meiotic cells. Intriguingly, this new work identified separable roles for 9-1-1 subunits, namely RAD9A-and HUS1-independent roles for RAD1. Based on these studies and the high-level expression of the paralogous proteins RAD9B and HUS1B in testis, we propose a model in which multiple alternative 9-1-1 clamps function during mammalian meiosis to ensure genome maintenance in the germline.

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Section: The 9-1-1 Complex Functions In Atr Activation and Dna Repairmentioning

confidence: 99%

Section: Rad9a and Hus1 Are Essential For A Subset Of Meiotic Recombimentioning

confidence: 99%

Clamping down on mammalian meiosis

et al. 2013

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“…His-tagged MutL and its derivative mutants were added to the GST-tagged RecA immobilized on glutathione-Sepharose 4B (Sigma, USA) and incubated for 1 h in 500 ll binding buffer (25 mM Tris-HCl, pH 7.5, 150 mM NaCl, 10 mM MgCl 2 , 10% glycerol, 1% Triton X-100, 0.5 mM DTT, 1% BSA) at 4°C. After centrifugation at 500g for 5 min, the pellets were washed five times with 500 ll binding buffer at 4°C and fractionated on a 10% SDS polyacrylamide gel [15]. Western blotting of His-tagged MutL and its derivative mutants was carried out with a primary antibody specific to polyHistidine (Monoclonal Anti-polyHistidine, Sigma, USA) and an anti-mouse-HRP conjugate (HRP-linked Anti-Mouse IgG, Sigma, USA) as a secondary antibody.…”

Section: In Vitro Pull-down Assaysmentioning

confidence: 99%

MutL associates with Escherichia coli RecA and inhibits its ATPase activity

Zhang

Zhou

et al. 2012

Archives of Biochemistry and Biophysics

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a b s t r a c t Different DNA repair systems are known to cooperate to deal with DNA damage. However, the regulatory role of the cross-talk between these pathways is unclear. Here, we have shown that MutL, an essential component of mismatch repair, is a RecA-interacting protein, and that its highly conserved N-terminal domain is sufficient for this interaction. Surface plasmon resonance and capillary electrophoresis analyses revealed that MutL has little effect on RecA-ssDNA filament formation, but dose down-regulate the ATPase activity of RecA. Our findings identify a new role for MutL, and suggest its regulatory role in homologous recombination.Ó 2011 Published by Elsevier Inc. IntroductionLiving organisms are continuously exposed to damaging agents from both internal and external environments. Cells have therefore evolved several repair systems to counteract potential threats to the stability of DNA and allow the accurate transmission of genetic information [1]. One such repair system is homologous recombination, a high fidelity DNA repair mechanism found in all forms of life that plays a very important role in the repair of double-stranded DNA breaks [2]. The Escherichia coli RecA protein plays an essential role in this repair pathway and has been used as a model system for studying DNA strand exchange, the central step of homologous recombination [3,4]. Like archaeal RadA and eukaryotic Rad51, bacterial RecA promotes an ATP-mediated strand exchange reaction by polymerizing on DNA and forming a helical filament [5].It is well known that there is cross-talk between different DNA repair pathways [6,7]. It has been suggested that there is cross-talk between homologous recombination and the mismatch repair system, since recombination between highly diverged sequences is differentially inhibited by the mismatch repair system [8,9]. MutL, a mismatch repair protein with DNA binding and ATPase activity, has also been reported to enhance MutS's inhibition of RecA-mediated homologous strand exchange between sequences which differ by 3% at the nucleotide level [10]. In mammalian cells, hMLH1, the homolog of MutL, also appears to monitor homologous recombination independent of mismatch repair [6]. However, the mechanism underlying the cross-talk between homologous recombination and the mismatch repair system, and in particular how MutL exerts its effect on homologous strand exchange, is unclear.Here we show that there is a physical interaction between MutL and the recombinase protein RecA, and that the highly conserved N-terminal domain of MutL is essential for this interaction. MutL down-regulates RecA ATPase activity, but does not affect RecA-ssDNA filament formation. Our results support the hypothesis that MutL may play a regulatory function in homologous recombination through its interaction with RecA. Materials and methods Protein purificationAll strains were grown routinely in Luria-Bertani (LB) media. Antibiotics were added when required at the following concentrations: ampicillin, 100 lg/ml; kanamycin, 60 lg/ml....

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“…Thus, the 9-1-1 complex may provide a platform for the assembly and function of the BER machinery (Balakrishnan et al, 2009;Lu et al, 2006). The 9-1-1 complex enhances mismatch repair via direct interaction with mismatch recognition proteins (MSH2/MSH3, MSH2/MSH6, and MLH1/PMS2) (Bai et al, 2010;He et al, 2008). Hus1 interacts with MSH2/MSH3 and MSH2/MSH6, but not with MLH1/PMS2 (Bai et al, 2010;He et al, 2008).…”

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confidence: 99%

“…The 9-1-1 complex enhances mismatch repair via direct interaction with mismatch recognition proteins (MSH2/MSH3, MSH2/MSH6, and MLH1/PMS2) (Bai et al, 2010;He et al, 2008). Hus1 interacts with MSH2/MSH3 and MSH2/MSH6, but not with MLH1/PMS2 (Bai et al, 2010;He et al, 2008). In the NER pathway, interactions between Saccharomyces cerevisiae Rad14 (hXPA homolog) and the checkpoint proteins ScDdc1 (hRad9 homolog) and ScMec3 (hHus1 homolog) have been demonstrated (Giannattasio et al, 2004).…”

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confidence: 99%