Human mismatch repair system balances mutation rates between strands by removing more mismatches from the lagging strand

Abstract

Mismatch repair (MMR) is one of the main systems maintaining fidelity of replication. Differences in correction of errors produced during replication of the leading and the lagging DNA strands were reported in yeast and in human cancers, but the causes of these differences remain unclear. Here, we analyze data on human cancers with somatic mutations in two of the major DNA polymerases, delta and epsilon, that replicate the genome. We show that these cancers demonstrate a substantial asymmetry of the mutations between the leading and the lagging strands. The direction of this asymmetry is the opposite between cancers with mutated polymerases delta and epsilon, consistent with the role of these polymerases in replication of the lagging and the leading strands in human cells, respectively. Moreover, the direction of strand asymmetry observed in cancers with mutated polymerase delta is similar to that observed in MMR-deficient cancers. Together, these data indicate that polymerase delta (possibly together with polymerase alpha) contributes more mismatches during replication than its leading-strand counterpart, polymerase epsilon; that most of these mismatches are repaired by the MMR system; and that MMR repairs about three times more mismatches produced in cells during lagging strand replication compared with the leading strand.

Figure 1.

Mutation frequencies in bMMRD cancers with subsequent mutations in Pol epsilon or Pol delta. Data for seven exomes of bMMRD cancer (five mutated Pol epsilon and two mutated Pol delta). Relative frequencies of single-nucleotide substitutions are shown irrespective of the strand; data from TCGA database for MMR-deficient (MSI) and MMR-proficient (MSS) samples without mutations in Pol epsilon and Pol delta are shown for comparison. N is the number of mutations observed in each sample.

Figure 2.

Replication asymmetry in bMMRD samples with mutations in replicative polymerases. (A) A schematic representation of estimation of the ratio of the mutation rates for complementary mutations. In the example shown, the frequency of the C-dT mismatch, resulting in the C → A mutation, is 1.5 times higher than the rate of the complementary G → T mutation when Pol epsilon is mutated and four times higher when Pol delta is mutated. Leftmost and rightmost bins together correspond to the 20% of the genome for which we can identify with the highest confidence that the reference strand is replicated primarily as lagging or leading, respectively. (B) The ratio of the mutation rates for complementary mutations (vertical axis) as a function of the propensity of the replication fork to replicate the reference strand as lagging or leading (horizontal axis). x-axis bins correspond to the percentiles of the replication timing derivative. Vertical bars represent 95% confidence intervals. Asterisks indicate significance of the deviation from one at the rightmost (or leftmost) bin. (*) P < 0.05; (**) P < 0.01; (***) P < 0.001. D corresponds to nucleotides A, G, or T. H corresponds to nucleotides A, C, or T. Note the logarithmic vertical axis.

Figure 3.

Comparison of mutational spectra of MMR-deficient and MMR-proficient cancers. Complementary mutation types were pooled. Data for MSI (n = 10) and MSS (n = 22) colon adenocarcinoma samples and uterine corpus endometrial carcinoma were pooled. Data for whole-genome sequencing. (A) Relative frequencies of the 96 mutation types (all possible mutation types in all possible tri-nucleotide contexts) in MSI and MSS cancers. (B) Ratio of the rates of each substitution in MSI and MSS cancers. Note the logarithmic vertical axis.

Figure 4.

Strand asymmetry of mutations. Panels show ratios of the mutation rates for complementary mutations in 10 MSI (three COAD and seven UCEC) and 22 MSS (eight COAD and 14 UCEC) cancers and in rare human polymorphisms. Whole-genome data used. Axes and notations are as in Figure 2B.

Figure 5.

Strand asymmetry of mutations in MSI cancers and in bMMRD cancers with mutated Pol epsilon and Pol delta cancers. (A) The asymmetry for complementary mutations in the 20% of the genome where replication asymmetry could be determined with the highest confidence (corresponding to bins 1 and 10 in Figs. 2B, 4). Error bars, 95% confidence intervals. (B) Model-estimated ratio of MMR effectiveness on the lagging and the leading strands.

Figure 6.

The schematic representation of MMR effectiveness during the leading and the lagging strand replication. While mismatches (red asterisks) are introduced more frequently during replication of the lagging strand by Pol delta, MMR corrects more mismatches on the lagging strand.