title: Rapid DNA double-strand breaks resulting from processing of Cr-DNA cross-links by both MutS dimers.
abstract: Mismatch repair (MMR) strongly enhances cyto- and genotoxicity of several chemotherapeutic agents and environmental carcinogens. DNA double-strand breaks (DSB) formed after two replication cycles play a major role in MMR-dependent cell death by DNA alkylating drugs. Here, we examined DNA damage detection and the mechanisms of the unusually rapid induction of DSB by MMR proteins in response to carcinogenic chromium(VI). We found that MSH2-MSH6 (MutSalpha) dimer effectively bound DNA probes containing ascorbate-Cr-DNA and cysteine-Cr-DNA cross-links. Binary Cr-DNA adducts, the most abundant form of Cr-DNA damage, were poor substrates for MSH2-MSH6, and their toxicity in cells was weak and MMR independent. Although not involved in the initial recognition of Cr-DNA damage, MSH2-MSH3 (MutSbeta) complex was essential for the induction of DSB, micronuclei, and apoptosis in human cells by chromate. In situ fractionation of Cr-treated cells revealed MSH6 and MSH3 chromatin foci that originated in late S phase and did not require replication of damaged DNA. Formation of MSH3 foci was MSH6 and MLH1 dependent, whereas MSH6 foci were unaffected by MSH3 status. DSB production was associated with progression of cells from S into G(2) phase and was completely blocked by the DNA synthesis inhibitor aphidicolin. Interestingly, chromosome 3 transfer into MSH3-null HCT116 cells activated an alternative, MSH3-like activity that restored dinucleotide repeat stability and sensitivity to chromate. Thus, sequential recruitment and unprecedented cooperation of MutSalpha and MutSbeta branches of MMR in processing of Cr-DNA cross-links is the main cause of DSB and chromosomal breakage at low and moderate Cr(VI) doses.
authors.affiliation: Department of Pathology, Laboratory Medicine, Brown University, Providence, Rhode Island 02912 , USA.
authors.forename: Mindy F
authors.initials: MF
authors.lastname: Reynolds
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Peterson-Roth
authors.forename: Ivan A
authors.initials: IA
authors.lastname: Bespalov
authors.forename: Tatiana
authors.initials: T
authors.lastname: Johnston
authors.forename: Volkan M
authors.initials: VM
authors.lastname: Gurel
authors.forename: Haley L
authors.initials: HL
authors.lastname: Menard
authors.forename: Anatoly
authors.initials: A
authors.lastname: Zhitkovich
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis; D002840:Chromates / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D002893:Chromosomes, Human, Pair 3; D018736:DNA Adducts / Q000737:chemistry / Q000378:metabolism*; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D045325:HCT116 Cells; D006367:HeLa Cells; D006801:Humans; D000070957:MutL Protein Homolog 1; D051719:MutS DNA Mismatch-Binding Protein / Q000378:metabolism*; D051718:MutS Homolog 2 Protein / Q000378:metabolism*; D009687:Nuclear Proteins / Q000096:biosynthesis; D017680:Potassium Compounds / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002840:Chromates; D018736:DNA Adducts; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; C094443:G-T mismatch-binding protein; C501486:MLH1 protein, human; D009687:Nuclear Proteins; D017680:Potassium Compounds; C027373:potassium chromate(VI); C497172:MSH2 protein, human; D000070957:MutL Protein Homolog 1; D051719:MutS DNA Mismatch-Binding Protein; D051718:MutS Homolog 2 Protein