High-accuracy lagging-strand DNA replication mediated by DNA polymerase dissociation

Abstract

The fidelity of DNA replication is a critical factor in the rate at which cells incur mutations. Due to the antiparallel orientation of the two chromosomal DNA strands, one strand (leading strand) is replicated in a mostly processive manner, while the other (lagging strand) is synthesized in short sections called Okazaki fragments. A fundamental question that remains to be answered is whether the two strands are copied with the same intrinsic fidelity. In most experimental systems, this question is difficult to answer, as the replication complex contains a different DNA polymerase for each strand, such as, for example, DNA polymerases δ and ε in eukaryotes. Here we have investigated this question in the bacterium , in which the replicase (DNA polymerase III holoenzyme) contains two copies of the same polymerase (Pol III, the gene product), and hence the two strands are copied by the same polymerase. Our in vivo mutagenesis data indicate that the two DNA strands are not copied with the same accuracy, and that, remarkably, the lagging strand has the highest fidelity. We postulate that this effect results from the greater dissociative character of the lagging-strand polymerase, which provides additional options for error removal. Our conclusion is strongly supported by results with antimutator polymerases characterized by increased dissociation rates.

New complexities of SOS-induced “untargeted” mutagenesis in Escherichia coli as revealed by mutation accumulation and whole-genome sequencing

“…In previous studies [12,22], we reported that in MMR-defective strains there is a 2-fold bias for A:T transitions to occur with the A on the LGST rather than on the LDST, and 2-fold bias for G:C transitions to occur with C on the LGST rather than on the LDST; these results were recently confirmed for transitions in the lacI gene [40]. We observed weaker but similar strand biases in MMR + strains [22].…”

New complexities of SOS-induced “untargeted” mutagenesis inEscherichia colias revealed by mutation accumulation and whole-genome sequencing

“…In previous studies [12,22], we reported that in MMR-defective strains there is a 2-fold bias for A:T transitions to occur with the A on the LGST rather than on the LDST, and 2-fold bias for G:C transitions to occur with C on the LGST rather than on the LDST; these results were recently confirmed for transitions in the lacI gene [40]. We observed weaker but similar strand biases in MMR + strains [22].…”

Overexpression of MutS impairs DNA mismatch repair and causes cell division defect inE.coli

“…In both cases, if the G/t mismatch goes unrepaired, the resultant GC to AT mutation would confer Lac + phenotype. It is shown previously that in E.coli , the fidelity of lagging strand replication is more as compared to that of leading strand 26,27 . Thus, during replication, E461G- lacZ -allele would bear far more G/t mismatches (correspondingly more chances of being Lac + ) at 1385 th position in native orientation as compared to that in invert orientation.…”