Structure and DNA-bridging activity of the essential Rec114–Mei4 trimer interface

Abstract

The DNA double-strand breaks (DSBs) that initiate meiotic recombination are formed by an evolutionarily conserved suite of factors that includes Rec114 and Mei4 (RM), which regulate DSB formation both spatially and temporally. In vivo, these proteins form large immunostaining foci that are integrated with higher-order chromosome structures. In vitro, they form a 2:1 heterotrimeric complex that binds cooperatively to DNA to form large, dynamic condensates. However, understanding of the atomic structures and dynamic DNA binding properties of RM complexes is lacking. Here, we report a structural model of a heterotrimeric complex of the C terminus of Rec114 with the N terminus of Mei4, supported by nuclear magnetic resonance experiments. This minimal complex, which lacks the predicted intrinsically disordered region of Rec114, is sufficient to bind DNA and form condensates. Single-molecule experiments reveal that the minimal complex can bridge two or more DNA duplexes and can generate force to condense DNA through long-range interactions. AlphaFold2 predicts similar structural models for RM orthologs across diverse taxa despite their low degree of sequence similarity. These findings provide insight into the conserved networks of protein–protein and protein–DNA interactions that enable condensate formation and promote formation of meiotic DSBs.

Evolutionary conservation of the structure and function of meiotic Rec114−Mei4 and Mer2 complexes

“…( F ) Model of Rec114 − Mei4 complex bound to two DNA duplexes. Residues implicated in DNA binding are shown as follows: R395, K396, K399, and R400 (4KR) in red ( Claeys Bouuaert et al 2021 ), and K403, K407, K417, and K424 in purple ( Liu et al 2023 ). The zoom only shows the position of the 4KR residues for clarity.…”

“…In addition, residues K403/K407 and K417/K424 also participate in DNA binding ( Liu et al 2023 ). Residues K417 and K424 of one monomer contribute in trans to the DNA-binding surface composed of residues R395, K396, K399, R400, K403, and K407 (6KR) of the second monomer ( Supplemental Fig.…”

Cryo-EM structure of the Spo11 core complex bound to DNA

“…However, the majority of particles again had just a single copy of the Spo11 core complex. It remains to be determined whether inclusion of other accessory proteins such as Rec114 and Mei4 is needed to allow dimer assembly and DNA cleavage in vitro [41][42][43][44] .…”

The molecular machinery of meiotic recombination

“…The recent protein structure prediction revolution, spearheaded by AlphaFold2 [ 145 ] has considerably lowered the barriers to high-resolution structural information necessary for point mutant design. From this, separation-of-function mutants can be used to study the details of meiotic recombination, as has been demonstrated recently [ 63 , 108 , 110 ]. At the time of writing, advanced prediction algorithms like AlphaFold2 do not yet possess the capability to model post-translational modifications, small molecule ligands, or nucleic acids.…”