Corentin Claeys Bouuaert

This work was led by my research technician Pascaline Liloku. Congratulations to her! I thank our collaborators, in particular the team of David Alsteens for help with AFM experiments, and Yann Sterckx for SAXS experiments. Finally, big thanks to the ERC and FNRS for funding!

Do we have any evidence to support this?

And does the DNA-binding activity of Spp1 have any functional consequences?

To find out, check out the paper!

We think that the binding of Spp1 to Mer2 occludes part of the DNA-binding interface of Mer2. However, the cost associated to Spp1 binding is compensated by a DNA-binding motif contributed by Spp1. We refer to this as an occlusion-compensation model.

So, how then do Mer2-DNA and Mer2-Spp1 interactions relate to each other? In other words, are the binding of DNA and Spp1 to Mer2 independent? Or are they competitive? Or in contrast are they cooperative?

We think there is yet another possibility.

So how does Spp1 bind DNA? Again AlphaFold proved helpful and pointed to a motif required for DNA binding.

Mutating this motif confirmed that, although full-length Spp1 does not bind DNA by itself, it binds DNA in the context of a complex with Mer2.

While full-length Spp1 does not bind DNA by itself, a truncation of Spp1 bound DNA quite efficiently. Thus, DNA-binding appears to be auto-inhibited in the context of the full-length protein.

That was surprise 3.

So how does Spp1 affect DNA binding and condensation?

We expected that DNA and Spp1 might compete for access to Mer2, but that doesn't seem to be the case. Instead Spp1 seems to somewhat stimulate DNA binding and condensation by Mer2.

Does Spp1 contribute to DNA binding directly?

Mer2 forms DNA-dependent condensates and effectively recruits Spp1.

Spp1 is essentially recruited as a client, but Spp1 does seem to stimulate condensation a little.

Now on two the second surprise: The Mer2 coiled coil domain that binds Spp1 is also involved in DNA binding.

We reconstituted Mer2-Spp1-DNA complexes and used AlphaFold to visualize what these complexes might look like.

(these models look pretty cool but are to be taken with a big grain of salt)

We verified that the previously-reported 4:2 stoichiometry was right. Indeed it was. Yet SAXS analysis all fit with the AlphaFold model.

There are several possible explanations: Our favorite is that the binding of Spp1 to Mer2 is allosterically regulated. (our arguments are in the paper).

However, there is an issue: based on the AlphaFold model, it is not clear why Mer2 and Spp1 would assemble a complex with a 4:2 stoichiometry, as had been shown previously. Mer2 is a homotetramer, and based on the model, it could accommodate 4 Spp1 subunits. That's our 1st surprise.

An mutagenesis analysis of the predicted interface provided evidence that supports the model.

We used AlphaFold to model the structure of the Mer2-Spp1 interaction domain, revealing a cool model with two Spp1 bound to a tetrameric Mer2 coiled coil.

We set out to characterize the interaction between Mer2 and Spp1.

We developed an approach based on atomic-force microscopy to measure single-molecule Spp1-Mer2 interactions, revealing a dynamic interaction.

Meiotic double-strand break (DSB) formation is tied to the loop-axis organization of meiotic chromosomes. DSB proteins are localized to the axes, and Spo11 cleaves DNA within loops. Spp1 connects the loop to the axes by binding H3K4me3 marks within loops and the DSB protein Mer2 on the axis.

Insights into the recruitment of the H3K4me3 reader Spp1 by the meiotic double-strand break protein Mer2 The formation of DNA double-strand breaks (DSBs) by Spo11 is tied to the loop-axis organization of meiotic chromosomes. Prior to DSB formation, chromatin loops marked by histone H3K4 trimethylation be...

✨New paper from the lab! ✨

We present a fun study aimed at characterizing the interaction between the H3K4me3 reader Spp1 and the meiotic double-strand break protein Mer2, and their relationship with DNA binding. We got some new insights and a few surprises.🧵

www.biorxiv.org/content/10.1...

Holliday junction–ZMM protein feedback enables meiotic crossover assurance - Nature Holliday junctions maintain chromosome synapsis to enable crossover assurance in budding yeast.

Holliday junction–ZMM feedback ensures meiotic crossover assurance in yeast! Great work from the Joao Matos lab!
👉 www.nature.com/articles/s41...

Protecting double Holliday junctions ensures crossing over during meiosis - Nature Conditional ablation experiments show that key components of the synaptonemal complex protect double Holliday junction recombination intermediates to ensure their resolution into crossover products, which are required for accurate chromosome segregation during meiosis.

Fantastic work coming out of Neil Hunter’s lab! www.nature.com/articles/s41...

RPA directly stimulates Mer3/HFM1 helicase processivity to ensure normal crossover formation in meiosis Meiotic crossover formation is critical for generating viable gametes and enhancing genetic diversity. The helicase Mer3 (HFM1 in humans) is a highly conserved factor essential for promoting crossover...

www.biorxiv.org/content/10.1...

We are hiring!
We are looking for a motivated and enthusiastic postdoc to study mammalian DSB repair using innovative genomic approaches.
Our lab is at the Curie Institute in Paris and offers vibrant scientific environnement and cutting edge platforms.
Please repost or apply here:
lnkd.in/eJ9q3QrR

We found a new asymmetry in the large-scale chromosome structure: sister chromatids are systematically shifted by hundreds of kb in the 5′→3′ direction of their inherited strands! The work was led by Flavia Corsi, in close collaboration with the Daniel Gerlich lab.
www.biorxiv.org/content/10.1...
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Thank you Valérie!

We’re happy to present a new preprint from the lab, where we identify new mechanisms that drive the recruitment of Mre11 to recombination sites during meiosis. Work led by star student Priyanka Priyadarshini with help from colleagues and funded by the ERC and FNRS.

www.biorxiv.org/content/10.1...

We’re happy to present a new preprint from the lab, where we identify new mechanisms that drive the recruitment of Mre11 to recombination sites during meiosis. Work led by star student Priyanka Priyadarshini with help from colleagues and funded by the ERC and FNRS.

www.biorxiv.org/content/10.1...

Diagram of two types of reproduction based on their meiosis. A. shows typical meiosis taking place in individuals with two copies of each chromosome (diploids) resulting in both male and female gametes carrying one copy of each chromosome. Fertilization restores the double copy for each chromosome. B. shows Canina meiosis in individuals carrying five copies of each chromosome (pentaploids) where male gametes only carry one copy of each chromosome and female gametes carry four copies of each chromosome. Fertilization restores the five copy for each chromosome.

Check out the Views & News piece that Joiselle Fernandes and I wrote for @nature.com on the recent work from @amarques.bsky.social and colleagues about one of the most bizarre ways to do meiosis rdcu.be/euabu.

Explained for non-experts!

Crossover patterning through condensation and coarsening of pro-crossover factors Nature Cell Biology - Zhang et al. provide evidence that, during meiosis, recombination proteins assemble into active droplets, the coarsening of which partially explains the phenomenon of...

Very happy (and frankly relieved) to see this paper finally out in peer-reviewed form. (Preprint was posted in August 2021, but this is the world we live in now). rdcu.be/erMk6

Maximizing meiotic crossover rates reveals the map of Crossover Potential - Nature Communications Meiotic crossovers enhance genetic diversity in sexually reproducing organisms. Here, the authors propose that the higher-order spatial organization of the meiotic chromosomes shapes sexual dimorphism...

Paper alert! #Meiosis4Ever

Maximizing meiotic crossover rates reveals the map of Crossover Potential

Juli Jing, Qiachao Lian and Stephanie Durand
www.nature.com/articles/s41...

We pushed meiotic crossover as much has we could, and had some surprises

A thread 👇

SPO11 dimers are sufficient to catalyse DNA double-strand breaks in vitro - Nature A biochemical system recapitulates the hallmarks of meiotic double-strand break formation, with mouse SPO11 catalysing break formation in the absence of any partners and remaining covalently atta...

Here is a link to the initial paper, in case you missed it.

www.nature.com/articles/s41...

In Vitro Reconstitution of SPO11-Mediated DNA Cleavage Sheds New Light on the Initiation of Meiotic Recombination | DNA and Cell Biology Three recent studies report the first biochemical reconstitution of DNA double-strand break (DSB) formation by SPO11, the topoisomerase-derived transesterase that initiates meiotic recombination in sexually reproducing organisms. A central conclusion of these studies is that SPO11 is sufficient to catalyze DSBs in vitro, but cleavage is limited by the poor propensity of SPO11 to dimerize, thereby providing an effective mechanism to prevent uncontrolled breaks. The studies yield new insights into the mechanism of DNA DSB formation and raise new questions regarding the functions of SPO11 partners, the impact of the DNA substrate, the coordination between cleavage events, and the reversibility of the reaction.

For your consideration, here is a short article where Cédric Oger and I reflect on our recent paper reporting the in vitro reconstitution of DNA cleavage by SPO11, and discuss some of the implications of this work.

The paper is available in open access.

www.liebertpub.com/doi/10.1089/...

Please see our latest paper on the role of EXO1 in meiosis: "EXO1 promotes the meiotic MLH1-MLH3 endonuclease through conserved interactions with MLH1, MSH4 and DNA". Congratulations to both first authors, Megha Roy and Aurore Sanchez and thanks to all our collaborators!