Kbiri, NadiaFernández Jiménez, NadiaDziegielewski, WojciechSáez-Zárate, EsperanzaPelé, AlexandreMata-Villanueva, AnaDluzewska, JuliaSantos Coloma, Juan LuisPradillo Orellana, MónicaZiolkowski, Piotr A2025-05-192025-05-192025-03-19Kbiri, N., Fernández-Jiménez, N., Dziegielewski, W., Sáez-Zárate, E., Pelé, A., Mata-Villanueva, A., Dluzewska, J., Santos, J. L., Pradillo, M., & Ziolkowski, P. A. (2025). Genetic dissection of MutL complexes in Arabidopsis meiosis. Nucleic Acids Research, 53(5). https://doi.org/10.1093/nar/gkaf1870305-104810.1093/nar/gkaf187https://hdl.handle.net/20.500.14352/120220Acknowledgements We thank Avi Levy (The Weizmann Institute, Israel) and Scott R. Poethig (University of Pennsylvania, US) for FTLs (CTLs); François Belzile (Université Laval, Canada) for sharing mlh1-1 and pms1-1, Raphaël Mercier (Max Planck Institute for Plant Breeding Research, Germany) for sharing mlh1-3; Franz Boideau for help with CoC analysis. B. Martín, M.C. Moreno, J. Barrios, and H. Korcz-Szatkowska for their excellent technical assistance. Funding This work was supported by the National Science Center, Poland (NCN) grants 2020/39/I/NZ2/02464 to P.A.Z. and 2019/35/N/NZ2/02933 to N.K. M.P. acknowledges the support of the Ministry of Science and Innovation of Spain (PID2020-118038GB-I00/AEI/10.13039/501100011033) and European Union (TED2021-131852B-100/ΑΕΙ/10.13039/501100011033/Unión Europea NextGeneration EU/PRTR). Funding to pay the Open Access publication charges for this article was provided by Initiative of Excellence-Research University at Adam Mickiewicz University, Poznan, PolandDuring meiosis, homologous chromosomes exchange genetic material through crossing over. The main crossover pathway relies on ZMM proteins, including ZIP4 and HEI10, and is typically resolved by the MLH1/MLH3 heterodimer, MutLγ. Our analysis shows that while MUS81 may partially compensate for MutLγ loss, its role remains uncertain. However, our multiple mutant analysis shows that MUS81 is unlikely to be the sole resolvase of ZMM-protected recombination intermediates when MutLγ is absent. Comparing genome-wide crossover maps of mlh1 mutants with ZMM-deficient mutants and lines with varying HEI10 levels reveals that crossover interference persists in mlh1 but is weakened. The significant crossover reduction in mlh1 also increases aneuploidy in offspring. The loss of MutLγ can be suppressed by eliminating the FANCM helicase. Combined with the lower-than-expected chiasma frequency, this suggests that in MutLγ absence, some ZMM-protected intermediates are ultimately resolved by DNA helicases and/or their complexes with Top3α. Elevated MLH1 or MLH3 expression moderately increases crossover frequency, while their misregulation drastically reduces crossover numbers and plant fertility, highlighting the importance for tight control of MLH1/MLH3 levels. By contrast, PMS1, a component of the MutLα endonuclease, appears uninvolved in crossing over. Together, these findings demonstrate the unique role of MutLγ in ZMM-dependent crossover regulation.engAttribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/journal article1362-4962https://doi.org/10.1093/nar/gkaf187open access575.113.1GenéticaBiología molecular (Biología)Fisiología vegetal (Biología)Biología celular (Biología)2409 Genética2415 Biología Molecular2407.02 Citogenética2417.14 Genética Vegetal