Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro

Dekang Liu, Jane H. Frederiksen, Sascha Emilie Liberti, Anne Lützen, Guido Keijzers, Javier Pena-Diaz, Lene Juel Rasmussen

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    Resumé

    DNA mismatch repair (MMR) is a highly-conserved DNA repair mechanism, whose primary role is to remove DNA replication errors preventing them from manifesting as mutations, thereby increasing the overall genome stability. Defects in MMR are associated with increased cancer risk in humans and other organisms. Here, we characterize the interaction between MMR and a proofreading-deficient allele of the human replicative DNA polymerase delta, PolδD316A;E318A, which has a higher capacity for strand displacement DNA synthesis than wild type Polδ. Human cell lines overexpressing PolδD316A;E318A display a mild mutator phenotype, while nuclear extracts of these cells exhibit reduced MMR activity in vitro, and these defects are complemented by overexpression or addition of exogenous human Exonuclease 1 (EXO1). By contrast, another proofreading-deficient mutant, PolδD515V, which has a weaker strand displacement activity, does not decrease the MMR activity as significantly as PolδD316A;E318A. In addition, PolδD515V does not increase the mutation frequency in MMR-proficient cells. Based on our findings, we propose that the proofreading activity restricts the strand displacement activity of Polδ in MMR. This contributes to maintain the nicks required for EXO1 entry, and in this manner ensures the dominance of the EXO1-dependent MMR pathway
    OriginalsprogEngelsk
    Artikelnummer28934474
    TidsskriftNucleic Acids Research
    Vol/bind45
    Udgave nummer16
    Sider (fra-til)9427-9440
    ISSN0305-1048
    DOI
    StatusUdgivet - 2017

    Citer dette

    Liu, D., Frederiksen, J. H., Liberti, S. E., Lützen, A., Keijzers, G., Pena-Diaz, J., & Rasmussen, L. J. (2017). Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro. Nucleic Acids Research, 45(16), 9427-9440. [28934474]. https://doi.org/10.1093/nar/gkx611
    Liu, Dekang ; Frederiksen, Jane H. ; Liberti, Sascha Emilie ; Lützen, Anne ; Keijzers, Guido ; Pena-Diaz, Javier ; Rasmussen, Lene Juel. / Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro. I: Nucleic Acids Research. 2017 ; Bind 45, Nr. 16. s. 9427-9440.
    @article{ec20a1956b804e1aa12cc22dfc149886,
    title = "Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro",
    abstract = "DNA mismatch repair (MMR) is a highly-conserved DNA repair mechanism, whose primary role is to remove DNA replication errors preventing them from manifesting as mutations, thereby increasing the overall genome stability. Defects in MMR are associated with increased cancer risk in humans and other organisms. Here, we characterize the interaction between MMR and a proofreading-deficient allele of the human replicative DNA polymerase delta, PolδD316A;E318A, which has a higher capacity for strand displacement DNA synthesis than wild type Polδ. Human cell lines overexpressing PolδD316A;E318A display a mild mutator phenotype, while nuclear extracts of these cells exhibit reduced MMR activity in vitro, and these defects are complemented by overexpression or addition of exogenous human Exonuclease 1 (EXO1). By contrast, another proofreading-deficient mutant, PolδD515V, which has a weaker strand displacement activity, does not decrease the MMR activity as significantly as PolδD316A;E318A. In addition, PolδD515V does not increase the mutation frequency in MMR-proficient cells. Based on our findings, we propose that the proofreading activity restricts the strand displacement activity of Polδ in MMR. This contributes to maintain the nicks required for EXO1 entry, and in this manner ensures the dominance of the EXO1-dependent MMR pathway",
    author = "Dekang Liu and Frederiksen, {Jane H.} and Liberti, {Sascha Emilie} and Anne L{\"u}tzen and Guido Keijzers and Javier Pena-Diaz and Rasmussen, {Lene Juel}",
    year = "2017",
    doi = "10.1093/nar/gkx611",
    language = "English",
    volume = "45",
    pages = "9427--9440",
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    Liu, D, Frederiksen, JH, Liberti, SE, Lützen, A, Keijzers, G, Pena-Diaz, J & Rasmussen, LJ 2017, 'Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro', Nucleic Acids Research, bind 45, nr. 16, 28934474, s. 9427-9440. https://doi.org/10.1093/nar/gkx611

    Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro. / Liu, Dekang; Frederiksen, Jane H.; Liberti, Sascha Emilie; Lützen, Anne; Keijzers, Guido; Pena-Diaz, Javier; Rasmussen, Lene Juel.

    I: Nucleic Acids Research, Bind 45, Nr. 16, 28934474, 2017, s. 9427-9440.

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    TY - JOUR

    T1 - Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro

    AU - Liu, Dekang

    AU - Frederiksen, Jane H.

    AU - Liberti, Sascha Emilie

    AU - Lützen, Anne

    AU - Keijzers, Guido

    AU - Pena-Diaz, Javier

    AU - Rasmussen, Lene Juel

    PY - 2017

    Y1 - 2017

    N2 - DNA mismatch repair (MMR) is a highly-conserved DNA repair mechanism, whose primary role is to remove DNA replication errors preventing them from manifesting as mutations, thereby increasing the overall genome stability. Defects in MMR are associated with increased cancer risk in humans and other organisms. Here, we characterize the interaction between MMR and a proofreading-deficient allele of the human replicative DNA polymerase delta, PolδD316A;E318A, which has a higher capacity for strand displacement DNA synthesis than wild type Polδ. Human cell lines overexpressing PolδD316A;E318A display a mild mutator phenotype, while nuclear extracts of these cells exhibit reduced MMR activity in vitro, and these defects are complemented by overexpression or addition of exogenous human Exonuclease 1 (EXO1). By contrast, another proofreading-deficient mutant, PolδD515V, which has a weaker strand displacement activity, does not decrease the MMR activity as significantly as PolδD316A;E318A. In addition, PolδD515V does not increase the mutation frequency in MMR-proficient cells. Based on our findings, we propose that the proofreading activity restricts the strand displacement activity of Polδ in MMR. This contributes to maintain the nicks required for EXO1 entry, and in this manner ensures the dominance of the EXO1-dependent MMR pathway

    AB - DNA mismatch repair (MMR) is a highly-conserved DNA repair mechanism, whose primary role is to remove DNA replication errors preventing them from manifesting as mutations, thereby increasing the overall genome stability. Defects in MMR are associated with increased cancer risk in humans and other organisms. Here, we characterize the interaction between MMR and a proofreading-deficient allele of the human replicative DNA polymerase delta, PolδD316A;E318A, which has a higher capacity for strand displacement DNA synthesis than wild type Polδ. Human cell lines overexpressing PolδD316A;E318A display a mild mutator phenotype, while nuclear extracts of these cells exhibit reduced MMR activity in vitro, and these defects are complemented by overexpression or addition of exogenous human Exonuclease 1 (EXO1). By contrast, another proofreading-deficient mutant, PolδD515V, which has a weaker strand displacement activity, does not decrease the MMR activity as significantly as PolδD316A;E318A. In addition, PolδD515V does not increase the mutation frequency in MMR-proficient cells. Based on our findings, we propose that the proofreading activity restricts the strand displacement activity of Polδ in MMR. This contributes to maintain the nicks required for EXO1 entry, and in this manner ensures the dominance of the EXO1-dependent MMR pathway

    U2 - 10.1093/nar/gkx611

    DO - 10.1093/nar/gkx611

    M3 - Journal article

    VL - 45

    SP - 9427

    EP - 9440

    JO - Nucleic Acids Research

    JF - Nucleic Acids Research

    SN - 0305-1048

    IS - 16

    M1 - 28934474

    ER -

    Liu D, Frederiksen JH, Liberti SE, Lützen A, Keijzers G, Pena-Diaz J et al. Human DNA polymerase delta double-mutant D316A;E318A interferes with DNA mismatch repair in vitro. Nucleic Acids Research. 2017;45(16):9427-9440. 28934474. https://doi.org/10.1093/nar/gkx611