Visualization of Eukaryotic DNA Mismatch Repair Reveals Distinct Recognition and Repair Intermediates

• Published in: Cell 2011-11, Vol.147 (5), p.1040-1053
• Main Authors: Hombauer, Hans, Campbell, Christopher S., Smith, Catherine E., Desai, Arshad, Kolodner, Richard D.
• Format: Article
• Language: English
• Subjects: active sites; Animals; base pair mismatch; DNA Mismatch Repair; DNA repair; DNA Repair Enzymes - metabolism; DNA Replication; DNA-Binding Proteins - metabolism; Exodeoxyribonucleases - metabolism; fluorescence; Humans; interphase; MutS Homolog 2 Protein - metabolism; Proliferating Cell Nuclear Antigen - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - metabolism
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/j.cell.2011.10.025

DNA mismatch repair (MMR) increases replication fidelity by eliminating mispaired bases resulting from replication errors. In Saccharomyces cerevisiae, mispairs are primarily detected by the Msh2-Msh6 complex and corrected following recruitment of the Mlh1-Pms1 complex. Here, we visualized functional fluorescent versions of Msh2-Msh6 and Mlh1-Pms1 in living cells. We found that the Msh2-Msh6 complex is an S phase component of replication centers independent of mispaired bases; this localized pool accounted for 10%–15% of MMR in wild-type cells but was essential for MMR in the absence of Exo1. Unexpectedly, Mlh1-Pms1 formed nuclear foci that, although dependent on Msh2-Msh6 for formation, rarely colocalized with Msh2-Msh6 replication-associated foci. Mlh1-Pms1 foci increased when the number of mispaired bases was increased; in contrast, Msh2-Msh6 foci were unaffected. These findings suggest the presence of replication machinery-coupled and -independent pathways for mispair recognition by Msh2-Msh6, which direct formation of superstoichiometric Mlh1-Pms1 foci that represent sites of active MMR. [Display omitted] ► S. cerevisiae Msh2-Msh6 is present in DNA replication factory foci ► Lagging-strand DNA mismatch repair (MMR) preferentially requires Exo1 ► Msh2-Msh6 foci-independent MMR is preferentially dependent on Exo1 ► Pms1 foci are active in MMR and contain substoichiometric amounts of Msh2-Msh6 In vivo imaging reveals that two mismatch repair protein complexes involved in mismatch repair have distinct functions, as one recognizes mispaired bases and one mediates active repair. The recognition complex is coupled to the DNA replication machinery, facilitating the detection of mismatches occurring during DNA synthesis.