Processing of O6-methylguanine by mismatch correction in human cell extracts

Curr Biol. 1996 Nov 1;6(11):1528-31. doi: 10.1016/s0960-9822(96)00758-0.


Human cell extracts perform an aberrant form of DNA synthesis on methylated plasmids [1], which represents processing of O6-methylguanine (O6-meG). Here, we show that extracts of colorectal carcinoma cells with defects in the mismatch repair proteins that normally correct replication errors do not carry out this synthesis. hMSH2-defective LoVo cell extracts (hMSH for human MutS homologue) performed O6-meG-dependent DNA synthesis only after the addition of the purified hMutS alpha mismatch recognition complex. Processing of O6-meG by mismatch correction requires PCNA and therefore probably DNA polymerase delta and/or epsilon. Mismatch repair-defective cells withstand O6-meG in their DNA [2], making them tolerant to methylating agents. Methylation-tolerant HeLaMR clones, with a mutator phenotype and a defect in either mismatch recognition or correction in vitro, also performed little O6-meG-dependent DNA synthesis. Assays of pairwise combinations of tolerant and colorectal carcinoma cell extracts identified hMLH1 as the missing mismatch repair function in a group of tolerant clones. The absence of processing by extracts of methylation-tolerant cells provides the first biochemical evidence that lethality of DNA O6-meG derives from its interaction with mismatch repair.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Extracts
  • DNA Repair*
  • DNA, Neoplasm / biosynthesis*
  • Guanine / analogs & derivatives*
  • Guanine / metabolism
  • HeLa Cells
  • Humans
  • Tumor Cells, Cultured


  • Cell Extracts
  • DNA, Neoplasm
  • Guanine
  • O-(6)-methylguanine