Biochemical characterisation of Mer3 helicase interactions and the protection of meiotic recombination intermediates

Altmannová, Veronika; Firlej, Magdalena; Müller, Franziska; Janning, Petra; Rauleder, Rahel; Rousová, Dorota; Schäffler, Andréas; Bange, Tanja; Weir, John R.

doi:10.1093/nar/gkad175

Cited by 6 publications

(7 citation statements)

References 71 publications

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“…Finally, streptavidin blotting revealed interesting information about the Mer3 helicase, whose function within the ZMM pathway is not fully understood (D uroc et al . 2017; A ltmannova et al . 2023).…”

Section: Discussionmentioning

confidence: 99%

Proximity labeling reveals new insights into the relationships between meiotic recombination proteins inS. cerevisiae

Voelkel-Meiman,

Poppel,

Liddle

et al. 2023

Preprint

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Several protein ensembles facilitate MutSγ crossover recombination and the associated process of synaptonemal complex (SC) assembly during meiosis, but the physical and functional relationships between the components involved remain obscure. We have employed proximity labeling as a phenotypic tool to discern functional relationships between meiotic recombination and SC proteins inS. cerevisiae, and to gain deeper insight into molecular deficits of crossover-defective mutants. We find that recombination initiation (Spo11) and the Mer3 helicase are dispensable for proximity labeling of the Zip3 E3 ligase by components of the ZZS ensemble (Zip2, Zip4 and Spo16) but are required for proximity labeling of Zip3 by Msh4, consistent with the possibility that MutSγ joins Zip3 only after a specific recombination intermediate has been generated. Proximity labeling analysis of crossover-defective zip1 mutants suggests a key shared defect is a failure to assemble an early recombination ensemble where ZZS can properly engage Zip3. We furthermore discovered that the abundance of Zip3 within the meiotic cell is uniquely dependent on the presence of Zip1, and that the post-translational modification of Zip3 is promoted by most MutSγ pathway proteins but countered by Zip1. Based on this and additional data, we propose a model whereby Zip1 stabilizes a functional, unmodified form of Zip3 until intermediate steps in recombination are complete. We also find that SC structural protein Ecm11 is proximity labeled by ZZS complex proteins in a Zip4-dependent manner, but by Zip3 and Msh4, at least in part, via a distinct pathway. Finally, streptavidin pulldowns followed by mass spectrometry on eleven different proximity labeling strains uncovers shared proximity targets of MutSγ-associated proteins, some with known meiotic functions and others not yet implicated in a meiotic activity, highlighting the potential power of proximity labeling as a discovery tool.

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Section: Discussionmentioning

confidence: 99%

Proximity labeling reveals new insights into the relationships between meiotic recombination proteins inS. cerevisiae

Voelkel-Meiman,

Poppel,

Liddle

et al. 2023

Preprint

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“…However, another MLH complex, Mlh1-Mlh2, has been implicated in regulating both crossover and noncrossover associated gene conversion tracts in conjunction with the Mer3 helicase. Recent studies proposed that Mer3-Mlh1-Mlh2 limits gene conversion through the inhibition of the Pif1, a 5’-3’ DNA helicase known to stimulate Polδ strand displacement through its interaction with the DNA polymerase processivity clamp PCNA (Abdullah et al 2004; Wilson et al 2013; Buzovetsky et al 2017; Duroc et al 2017; Altmannova et al 2023). In support of this model, Vernekar et al (2021) found that Mer3 interacts with both Pif1 and Rfc1 in vivo and that the Mer3-Mlh1-Mlh2 complex blocks Polδ-mediated D-loop extension in vitro .…”

Section: Discussionmentioning

confidence: 99%

The Dmc1 recombinase physically interacts with and promotes the meiotic crossover functions of the Mlh1-Mlh3 endonuclease

Pannafino,

Chen,

Mithani

et al. 2023

Preprint

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The accurate segregation of homologous chromosomes during the Meiosis I reductional division in most sexually reproducing eukaryotes requires crossing over between homologs. In baker’s yeast approximately 80 percent of meiotic crossovers result from Mlh1-Mlh3 and Exo1 acting to resolve double-Holliday junction (dHJ) intermediates in a biased manner. Little is known about how Mlh1-Mlh3 is recruited to recombination intermediates and whether it interacts with other meiotic factors prior to its role in crossover resolution. We performed a haploinsufficiency screen in baker’s yeast to identify novel genetic interactors with Mlh1-Mlh3 using sensitizedmlh3alleles that disrupt the stability of the Mlh1-Mlh3 complex and confer defects in mismatch repair but do not disrupt meiotic crossing over. We identified several genetic interactions betweenMLH3andDMC1,the recombinase responsible for recombination between homologous chromosomes during meiosis. We then showed that Mlh3 physically interacts with Dmc1in vitroand at times in meiotic prophase when Dmc1 acts as a recombinase. Interestingly, restrictingMLH3expression to roughly the time of crossover resolution resulted in amlh3null-like phenotype for crossing over. Our data are consistent with a model in which Dmc1 nucleates a polymer of Mlh1-Mlh3 to promote crossing over.

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“…Mer3 is a helicase with many extra domains beyond its helicase core. It is most closely related to the spliceosomal RNA helicase Brr2 [ 11 , 125 ]. The helicase activity of Mer3 has been previously suggested to expand nascent D-loops, thus stabilising them.…”

Section: Zmm Proteins In Crossover Formationmentioning

confidence: 99%

“…The helicase activity of Mer3 has been previously suggested to expand nascent D-loops, thus stabilising them. Indeed, in vitro Mer3 clearly has a strong preference for D-loop DNA [ 11 , 126 ]. The preference for D-loop DNA binding suggests that Mer3 may bind to early recombination intermediates.…”

Section: Zmm Proteins In Crossover Formationmentioning

confidence: 99%

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The molecular machinery of meiotic recombination

Chen,

Weir

2024

Biochemical Society Transactions

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Meiotic recombination, a cornerstone of eukaryotic diversity and individual genetic identity, is essential for the creation of physical linkages between homologous chromosomes, facilitating their faithful segregation during meiosis I. This process requires that germ cells generate controlled DNA lesions within their own genome that are subsequently repaired in a specialised manner. Repair of these DNA breaks involves the modulation of existing homologous recombination repair pathways to generate crossovers between homologous chromosomes. Decades of genetic and cytological studies have identified a multitude of factors that are involved in meiotic recombination. Recent work has started to provide additional mechanistic insights into how these factors interact with one another, with DNA, and provide the molecular outcomes required for a successful meiosis. Here, we provide a review of the recent developments with a focus on protein structures and protein–protein interactions.

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Biochemical characterisation of Mer3 helicase interactions and the protection of meiotic recombination intermediates

Cited by 6 publications

References 71 publications

Proximity labeling reveals new insights into the relationships between meiotic recombination proteins inS. cerevisiae

Proximity labeling reveals new insights into the relationships between meiotic recombination proteins inS. cerevisiae

The Dmc1 recombinase physically interacts with and promotes the meiotic crossover functions of the Mlh1-Mlh3 endonuclease

The molecular machinery of meiotic recombination

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