269: Mlh1–Pms1 endonuclease creates single-strand gaps to excise mispairs in S. cerevisiae MMR

Palacio T et al., Proc. Natl. Acad. Sci. U.S.A. 2025.122:e2528670122 - Reconstituted Saccharomyces cerevisiae mismatch repair shows the Mlh1–Pms1 endonuclease directly generates single-strand gaps to excise mispairs independent of Exo1 and Rad27. Key terms: Mlh1-Pms1, mismatch repair, single-strand gaps, Exo1, APOBEC3A. Study Highlights:We reconstituted Saccharomyces cerevisiae mismatch repair with purified Msh2–Msh6 or Msh2–Msh3, Mlh1–Pms1, PCNA, RFC, RPA, and DNA polymerases and analyzed products by restriction mapping, Mung Bean nuclease, electron microscopy, and APOBEC3A deamination. The Mlh1–Pms1 endonuclease, activated by RFC-loaded PCNA and Mn2+, generates strand-specific single-strand gaps on the preexisting nicked strand. Electron microscopy and deamination mapping revealed a broad distribution of gap sizes with a peak around 128 ± 17 nucleotides and most gaps under 500 nucleotides. These gaps can be filled by DNA Polε or low levels of DNA Polδ, providing an Exo1- and Rad27-independent route for mispair excision and explaining redundancy among excision pathways. Conclusion:Mlh1–Pms1 catalyzes strand-specific single-strand gap formation that mediates Exo1- and Rad27-independent mispair excision in reconstituted S. cerevisiae mismatch repair. QC:This episode was checked against the original article PDF and publication metadata for the episode release published on 2026-01-24. QC Scope:- article metadata and core scientific claims from the narration- excludes analogies, intro/outro, and music- transcript coverage: Audited sections: Exo1/Rad27 independence, MLH1-Pms1 endonuclease role, Mn2+ activation, gap formation and sizes, EM evidence, APOBEC3A gap mapping, and the proposed three-pathway MMR model.- transcript topics: Background on mismatch repair and Exo1/Rad27 roles; Exo1- and Rad27-independent MMR reconstitution; MLH1-Pms1 endonuclease activity and Mn2+ activation; Formation and types of single-stranded gaps; Electron microscopy evidence of gap sizes; APOBEC3A mapping of excision gaps QC Summary:- factual score: 10/10- metadata score: 10/10- supported core claims: 7- claims flagged for review: 0- metadata checks passed: 4- metadata issues found: 0 Metadata Audited:- article_doi- article_title- article_journal- license Factual Items Audited:- Combined Exo1- and Rad27-independent repair accounts for a small fraction (5–13%) of MMR when Exo1/Rad27 pathways are deleted, with Exo1/Rad27 contributing the remainder via redund- MLH1–Pms1 endonuclease can drive mispair excision independently of Exo1 and Rad27.- MLH1–Pms1–mediated repair generates strand-specific single-stranded gaps on the nicked strand.- Gap sizes are broadly distributed with a peak at 128 ± 17 nucleotides; most gaps are under 500 nucleotides.- Gaps can be filled by DNA Polε or low levels of DNA Polδ, enabling Exo1- and Rad27-independent repair.- Mn2+ activates MLH1–Pms1 endonuclease; Mg2+ alone does not support robust activity. QC result: Pass. Chapters (00:00:00) - Base by Bass(00:00:29) - Who is the real repairman of DNA?(00:02:39) - The spell-checker for our DNA(00:04:23) - The ghost mechanic of DNA repair(00:07:02) - The secret to DNA repair(00:09:48) - How DNA repair works: The 3 pathways(00:13:43) - How to Stop cancer with a single drug