DNA mismatch repair mediated by Mlh1-Pms1 endonuclease-catalyzed mispair excision

Area of Science:

• Molecular Biology
• Genetics
• DNA Repair Mechanisms

Background:

• Eukaryotic DNA mismatch repair (MMR) relies on multiple excision pathways.
• Key pathways involve exonuclease 1 (Exo1) and flap endonuclease Rad27 (FEN1) with DNA polymerase δ.
• Inactivating only Exo1 and Rad27 results in a minor MMR defect, suggesting alternative mechanisms.

Purpose of the Study:

• To investigate the role of Mlh1-Pms1 in nicked-strand-specific MMR.
• To reconstitute MMR in vitro under conditions lacking Exo1 and Rad27.
• To understand the interdependence and redundancy of MMR excision pathways.

Main Methods:

• Reconstitution of nicked-strand-specific MMR using purified Saccharomyces cerevisiae proteins.
• In vitro assays were performed in the absence of Exo1, Rad27, or strand-displacement synthesis.
• Investigated the requirement for Mlh1-Pms1 endonuclease activity, its activation by RFC/PCNA, and recruitment by Msh2-Msh6/Msh2-Msh3.

Main Results:

• Mlh1-Pms1 endonuclease activity, activated by RFC and PCNA, and recruited by Msh2-Msh6/Msh2-Msh3, was essential for MMR.
• MMR was mediated by Mlh1-Pms1-dependent excision, creating single-strand DNA gaps of variable sizes.
• These findings support genetic data on the redundancy of Exo1, Rad27, and Mlh1-Pms1 in MMR.

Conclusions:

• The Mlh1-Pms1 endonuclease can independently mediate nicked-strand-specific MMR.
• This demonstrates functional redundancy among Exo1, Rad27, and Mlh1-Pms1 excision pathways.
• MMR exhibits flexibility, utilizing alternative excision mechanisms to maintain genomic integrity.