Multiple mechanisms limit meiotic crossovers: TOP3α and two BLM homologs antagonize crossovers in parallel to FANCM

Abstract

Meiotic crossovers (COs) have two important roles, shuffling genetic information and ensuring proper chromosome segregation. Despite their importance and a large excess of precursors (i.e., DNA doublestrand breaks, DSBs), the number of COs is tightly regulated, typically one to three per chromosome pair. The mechanisms ensuring that most DSBs are repaired as non-COs and the evolutionary forces imposing this constraint are poorly understood. Here we identified Topoisomerase3α (TOP3α) and the RECQ4 helicases-the Arabidopsis slow growth suppressor 1 (Sgs1)/Bloom syndrome protein (BLM) homologs-as major barriers to meiotic CO formation. First, the characterization of a specific TOP3α mutant allele revealed that, in addition to its role in DNA repair, this topoisomerase antagonizes CO formation. Further, we found that RECQ4A and RECQ4B constitute the strongest meiotic anti-CO activity identified to date, their concomitant depletion leading to a sixfold increase in CO frequency. In both top3α and recq4ab mutants, DSB number is unaffected, and extra COs arise from a normally minor pathway. Finally, both TOP3α and RECQ4A/B act independently of the previously identified anti-CO Fanconi anemia of complementation group M (FANCM) helicase. This finding shows that several parallel pathways actively limit CO formation and suggests that the RECQA/B and FANCM helicases prevent COs by processing different substrates. Despite a ninefold increase in CO frequency, chromosome segregation was unaffected. This finding supports the idea that CO number is restricted not because of mechanical constraints but likely because of the long-term costs of recombination. Furthermore, this work demonstrates how manipulating a few genes holds great promise for increasing recombination frequency in plant-breeding programs.eiotic homologous recombination is initiated by the formation of DNA double-strand breaks (DSBs). DSBs are resected to form 3′ ssDNA overhangs which invade the intact homologous chromosome, producing DNA joint molecules (JMs). These JMs can be differentially processed to produce crossovers (COs) or non-COs (NCOs). In Arabidopsis thaliana, mammals, and budding yeast, two pathways of CO formation exist. The major pathway depends on the ZMM proteins (for Zip1-4, Msh4/5, and Mer3) in addition to MutL homolog 1 (MLH1) and MuL homolog 3 (MLH3) and produces interfering COs, so that one CO prevents the formation of another nearby (1). The second, pathway, producing noninterfering COs, depends on structure-specific endonucleases including MUS81 (1). These pro-CO pathways compete with anti-CO pathways, resulting in a minor portion of DSBs becoming COs; for instance, it is estimated that COs represent only 10% and 5% of DSBs in mouse and in Arabidopsis, respectively (2). Three helicases with meiotic anti-CO activities have been identified in different species: the Bloom syndrome (BLM) homolog, small growth suppressor 1 (Sgs1) in Saccharomyces cerevisiae (3, 4); regulator of telomere elongation helicase1 (RTEL-1) in Caenorhabditis elegans (5)...

Functions and regulation of the multitasking FANCM family of DNA motor proteins

“…Nonoverlapping DNA repair roles between Mph1 and either Sgs1 or Srs2 lead to synthetic sickness or lethality of double mutants upon DNA damage occurrence (St Onge et al 2007;Chen et al 2009;Panico et al 2010). Similarly, Mph1 and STR orthologs in fission yeast and A. thaliana also have independent functions in crossover control and/or DNA repair (Sun et al 2008;Knoll et al 2012;Seguela-Arnaud et al 2015). In D. melanogaster, FANCM and BTR appear to be epistatic for crossover control but additive for DNA repair (Kuo et al 2014).…”

The Structure-Specific Endonucleases MUS81 and SEND1 Are Essential for Telomere Stability in Arabidopsis

“…Thus, RECQ4B does not play a detectable role in somatic DNA repair. However, both RECQ4A and RECQ4B helicases have anticrossover activity in meiosis, with their simultaneous depletion inducing a 6-fold increase in CO frequency (Séguéla-Arnaud et al, 2015).…”

DNA methylation epigenetically silences crossover hot spots and controls chromosomal domains of meiotic recombination in Arabidopsis

“…The FANCM helicase acts with MHF1 and MHF2 to promote formation of noncrossovers, in the absence of which most strand invasion events enter a noninterfering crossover repair pathway (Crismani et al 2012;Knoll et al 2012;Girard et al 2014). FIDGITIN, RECQ4A, RECQ4B, and TOPOISOMERASE3α have been identified as additional anti-crossover factors in Arabidopsis (Girard et al 2015;Séguéla-Arnaud et al 2015).…”