Cadmium is a mutagen that acts by inhibiting mismatch repair

Abstract

Most errors that arise during DNA replication can be corrected by DNA polymerase proofreading or by post-replication mismatch repair (MMR). Inactivation of both mutation-avoidance systems results in extremely high mutability that can lead to error catastrophe. High mutability and the likelihood of cancer can be caused by mutations and epigenetic changes that reduce MMR. Hypermutability can also be caused by external factors that directly inhibit MMR. Identifying such factors has important implications for understanding the role of the environment in genome stability. We found that chronic exposure of yeast to environmentally relevant concentrations of cadmium, a known human carcinogen, can result in extreme hypermutability. The mutation specificity along with responses in proofreading-deficient and MMR-deficient mutants indicate that cadmium reduces the capacity for MMR of small misalignments and base-base mismatches. In extracts of human cells, cadmium inhibited at least one step leading to mismatch removal. Together, our data show that a high level of genetic instability can result from environmental impediment of a mutation-avoidance system.

Figure 1

The impact of CdCl₂ on mutation rates and viability in yeast. Yeast strains are described in Supplementary Methods online. Rates, frequencies and 95% confidence intervals are provided in Supplementary Table 1 online. Frameshift mutation reporters with homonucleotide runs in the CG379 background included lys2-A₁₄ (A14 run that reverts by -1 frameshifts), lys2-A₁₂ (A12 run that reverts by +1 frameshifts), lys2-A₁₀ (A10 run that reverts by -1 frameshifts) and his7-2 (A7 run that reverts by +1 frameshifts). Homonucleotide run reporters containing 10 bases of A, T, G or C at the same position in LYS2 that revert by -1 frameshifts were also used in the SJR background. The CAN1 forward mutations arise by frameshifts, base substitutions and gross rearrangements. Yeast strains and mutation reporters are indicated on the corresponding panels. (a) Mutagenesis in homonucleotide runs of wild-type strains. WT, wild-type strains of CG379 background; SJR, wild-type strains of SJR background. The SJR strains were more resistant to killing by cadmium and required higher doses to reach a mutagenesis plateau. The rates of his7-2 reversion were determined in the wild-type strain carrying lys2-A₁₄. They were similar to rates of mutation in the his7-2 reporter determined in the isogenic wild-type strains carrying lys2-A₁₂ and lys2-A₁₀ (Supplementary Table 1 online). (b) Mutagenesis in MMR-deficient strains. Mutation rates for wild-type lys2-A₁₄ are the same as in a. (c) Viability of wild-type and proofreading-deficient strains. All strains are in CG379 background. 1n, WT indicates a wild-type strain with lys2-A₁₄ reporter, which was used to create all other strains shown on this panel. 1n, haploid; 2n, diploid. (d) Mutagenesis in proofreading-deficient haploid (1n) and diploid (2n) strains. Strains are the same as in c. Only pol3-01 diploids, but not haploids, were used to avoid the lethal effect of cadmium. Because can1 mutations are recessive, they cannot be assessed in pol3-01 diploids; therefore, can1 rates were determined only for the haploid Pol ε proofreading-deficient mutant pol2-4. The data for wild-type haploid strains are the same as in panels a and b.

Figure 2

Inhibition of in vitro human strand-specific DNA MMR in a repair-proficient cell extract by cadmium. Data are the average of three independent experiments. Error bars represent standard deviation. The average percent reduction in repair efficiency, taking the amount of repair in the absence of cadmium as 100%, is shown in parentheses above each bar.