DNA mismatch repair-dependent response to fluoropyrimidine-generated damage

Mark Meyers, Mark W. Wagner, Anthony Mazurek, Christoph Schmutte, Richard Fishel, David A. Boothman

Research output: Contribution to journalArticle

107 Scopus citations

Abstract

Previous studies from our laboratory indicated that expression of the MLH1 DNA mismatch repair (MMR) gene was necessary to restore cytotoxicity and an efficient G2 arrest in HCT116 human colon cancer cells, as well as Mlh1-/- murine embryonic fibroblasts, after treatment with 5-fluoro-2′-deoxyuridine (FdUrd). Here, we show that an identical phenomenon occurred when expression of MSH2, the other major MMR gene, was restored in HEC59 human endometrial carcinoma cells or was present in adenovirus E1A-immortalized Msh2+/+ (compared with isogenic Msh2-/-) murine embryonic stem cells. Because MMR status had little effect on cellular responses (i.e. G2 arrest and lethality) to the thymidylate synthase inhibitor, Tomudex, and a greater level of [3H]FdUrd incorporation into DNA was found in MMR-deficient cells, we concluded that the differential FdUrd cytotoxicity between MMR-competent and MMR-deficient cells was mediated at the level of DNA incorporation. Analyses of ATPase activation suggested that the hMSH2-hMSH6 heterodimer only recognized FdUrd moieties (as the base 5-fluorouracil (FU) in DNA) when mispaired with guanine, but not paired with adenine. Furthermore, analyses of incorporated FdUrd using methyl-CpG-binding domain 4 glycosylase indicated that there was more misincorporated FU:Gua in the DNA of MMR-deficient HCT116 cells. Our data provide the first demonstration that MMR specifically detects FU:Gua (in the first round of DNA replication), signaling a sustained G2 arrest and lethality.

Original languageEnglish (US)
Pages (from-to)5516-5526
Number of pages11
JournalJournal of Biological Chemistry
Volume280
Issue number7
DOIs
StatePublished - Feb 18 2005

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Meyers, M., Wagner, M. W., Mazurek, A., Schmutte, C., Fishel, R., & Boothman, D. A. (2005). DNA mismatch repair-dependent response to fluoropyrimidine-generated damage. Journal of Biological Chemistry, 280(7), 5516-5526. https://doi.org/10.1074/jbc.M412105200