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.