HO Endonuclease-Initiated Recombination in Yeast Meiosis Fails To Promote Homologous Centromere Pairing and Is Not Constrained To Utilize the Dmc1 Recombinase

Abstract

Crossover recombination during meiosis is accompanied by a dramatic chromosome reorganization. In Saccharomyces cerevisiae, the onset of meiotic recombination by the Spo11 transesterase leads to stable pairwise associations between previously unassociated homologous centromeres followed by the intimate alignment of homologous axes via synaptonemal complex (SC) assembly. However, the molecular relationship between recombination and global meiotic chromosome reorganization remains poorly understood. In budding yeast, one question is why SC assembly initiates earliest at centromere regions while the DNA double strand breaks (DSBs) that initiate recombination occur genome-wide. We targeted the site-specific HO endonuclease to various positions on S. cerevisiae’s longest chromosome in order to ask whether a meiotic DSB’s proximity to the centromere influences its capacity to promote homologous centromere pairing and SC assembly. We show that repair of an HO-mediated DSB does not promote homologous centromere pairing nor any extent of SC assembly in spo11 meiotic nuclei, regardless of its proximity to the centromere. DSBs induced en masse by phleomycin exposure likewise do not promote homologous centromere pairing nor robust SC assembly. Interestingly, in contrast to Spo11, HO-initiated interhomolog recombination is not affected by loss of the meiotic kinase, Mek1, and is not constrained to use the meiosis-specific Dmc1 recombinase. These results strengthen the previously proposed idea that (at least some) Spo11 DSBs may be specialized in activating mechanisms that both 1) reinforce homologous chromosome alignment via homologous centromere pairing and SC assembly, and 2) establish Dmc1 as the primary strand exchange enzyme.

Rad51 and Dmc1 have similar tolerance for mismatches in yeast meiosis

“…where Dmc1 is the recombinase), suggesting that Msh2 enforced heteroduplex rejection, even though there is no 3’ end nonhomology. These results suggests that wild type meiotic cells, even acting at an HO-induced DSB not created by Spo11 and thus not necessarily in association with the entire apparatus of the meiotic recombination machinery (79) interacts differently with Msh2 than does Rad51.…”

“…We conclude that, in the context of the intact recombination machinery, Dmc1-mediated recombination is not intrinsically more tolerant of mismatches than that driven by Rad51, at least during ectopic BIR in meiosis, using a site-specific DSB. Although meiotic recombination outcomes of SPO13::HO-initiated events are distinctly meiosis-like (69), it is distinctly possible that some parts of the meiosis machinery are intrinsically linked to the elaborate protein assemblies associated with the creation of Spo11-mediated DSBs (79). The ability to create a specific Spo11 hotspot in a normally “cold” region of the genome (83) may help elucidate these questions.…”

Meiosis in budding yeast

“…At the same time, hop1 mutants display genome-wide shifts in loop structure ( Schalbetter et al 2019 ) and loss of either Red1 or Hop1 results in abnormally diffuse mid-meiotic prophase chromosome morphology, indicating a role for these proteins in chromosome compaction ( Nag et al 1995 ; Yu and Koshland 2003 ). Red1 and Hop1 may affect higher-order chromatin folding via their critical functions in meiotic DSB formation, as a DSB-deficient spo11 mutant displays chromosome individualization defects similar to red1 and hop1 mutants ( Mao-Draayer et al 1996 ; Smith and Roeder 1997 ; Klein et al 1999 ; Macqueen and Roeder 2009 ; Yisehak and MacQueen 2018 ).…”

“…DSB formation and strand exchange ( Loidl et al 1994 ; Weiner and Kleckner 1994 ; Peoples et al 2002 ; Peoples-Holst and Burgess 2005 ; Lui et al 2006 ). Yet, homolog recognition does not appear to be solely underpinned by strand exchange, as mutants lacking both Rad51 and Dmc1 exhibit a substantial level of homolog pairing compared to the low level observed in a spo11 mutant ( Tsubouchi and Roeder 2003 ; Yisehak and Macqueen 2018 ).…”

A Grave Affair

“…Cells without SPO13 experience a failure to mono-orient sister kinetochores in meiosis I and an inability to protect cohesin around the centromere [91]. Although we do not know the exact role of Spo13 in cohesin protection, this function does not appear to be an essential role during meiosis as the diploid dyads that result from a recombination defective mutant (spo13∆ spo11∆) are 89% viable suggesting meiosis II chromosome segregation has occurred properly [114]. It is intriguing that the Spo13 protein appears to be genetically interacting with the fpr3∆ zip3∆ mutant, so we note the possibility that Spo13 itself is detrimental to meiosis when Fpr3 and Zip3 are missing.…”