ANKRD31 regulates spatiotemporal patterning of meiotic recombination initiation and ensures recombination between heterologous sex chromosomes in mice

Abstract

Orderly segregation of chromosomes during meiosis requires that crossovers form between homologous chromosomes by recombination. Programmed DNA double-strand breaks (DSBs) initiate meiotic recombination. We identify ANKRD31 as a critical component of complexes of DSB-promoting proteins which assemble on meiotic chromosome axes. Genome-wide, ANKRD31 deficiency causes delayed recombination initiation. In addition, loss of ANKRD31 alters DSB distribution owing to reduced selectivity for sites that normally attract DSBs.Strikingly, ANKRD31 deficiency also abolishes uniquely high rates of recombination that normally characterize pseudoautosomal regions (PARs) of X and Y chromosomes.Consequently, sex chromosomes do not form crossovers leading to chromosome segregation failure in ANKRD31-deficient spermatocytes. These defects are accompanied by a genome-wide delay in assembling DSB-promoting proteins on axes and a loss of a specialized PAR-axis domain that is highly enriched for DSB-promoting proteins. Thus, we propose a model for spatiotemporal patterning of recombination by ANKRD31-dependent control of axis-associated complexes of DSB-promoting proteins. Highlights (85 characters max)Temporal and spatial patterning of recombination are regulated by ANKRD31 Selective use of PRDM9 binding sites as DSB hotspots requires ANKRD31 Enrichment of pro-DSB factors in the PAR requires ANKRD31 but not IHO1 Recombination in the PAR critically depends on ANKRD31 are consistent with unpublished data of Acquaviva, Jasin & Keeney (personal communication).Acquaviva et al. observed ANKRD31 aggregates on PARs, and identified chromosome 4, 9 and 13 as autosomes whose non-centromeric ends carry arrays of PAR-like sequences that associate with ANKRD31 aggregates. Distinct molecular requirements for ANKRD31 aggregates and ANKRD31 fociGiven their distinct behaviour in synapsed regions, foci and aggregates of ANKRD31/MEI4/REC114 might represent qualitatively different protein complexes with distinct underlying molecular requirements. To test this possibility we compared localization of ANKRD31, REC114 and MEI4 in Mei4 -/-, Rec114 -/and Iho1 -/spermatocytes ( Figure 2J and S3E-I). The numbers of REC114 and MEI4 foci are strongly reduced in Mei4 -/and Rec114 -/mice, respectively (Kumar et al., 2018). In addition, we found that both focus and aggregate formation of ANKRD31, REC114 and MEI4 were each disrupted in Mei4 -/and Rec114 -/spermatocytes ( Figure S3E-G and Table S3). Remarkably, only the formation of ANKRD31 ( Figure 2I), MEI4 and REC114 foci (Stanzione et al., 2016), but not aggregates, were disrupted in Iho1 -/spermatocytes ( Figure 2J). ANKRD31 aggregates formed efficiently in Iho1 -/spermatocytes; median numbers of ANKRD31 aggregates were four in both wild type (n=62) and Iho1 -/-(n=57) spermatocytes in zygotene. These aggregates always colocalized with aggregates of MEI4 (n=100) and REC114 (n=100) in Iho1 -/spermatocytes ( Figure 2J and S3H).ANKRD31 aggregates also colocalized with PAR FISH signals in late zygotene-like Iho1 -/spe...

Ensuring meiotic DNA break formation in the mouse pseudoautosomal region

“…MEI4 is thus essential for assembly of RMMAI foci genome wide, while ANKRD31 contributes genome-wide but is more specifically essential for high-level enrichment at mo-2 regions. These results agree with findings of Tóth and colleagues, who further observed that REC114 but not IHO1 is essential for formation of blobs of the other RMMAI proteins (31).…”

“…We found that the PAR was highly enriched for DSB-promoting factors REC114, MEI4, MEI1, and IHO1 [proteins essential for genomewide DSB formation (22-24, 29)] as well as ANKRD31, a REC114 partner essential for PAR DSB formation (30,31). All five proteins (hereafter RMMAI for simplicity) colocalized in several bright, irregular "blobs" for most of prophase I (Fig.…”

“…5D). [Ankrd31 -/mutants form substantially fewer RPA2 foci and other cytological DSB markers at leptonema and early zygonema, but not from mid-zygonema on (30,31).] As a consequence, the X and Y chromosomes were paired in only 6% of mid-pachytene spermatocytes ( Fig.…”

“…As a consequence, the X and Y chromosomes were paired in only 6% of mid-pachytene spermatocytes ( Fig. 5F), whereas most Ankrd31 -/cells pair and synapse all autosomes (30,31).…”

“…To test more directly whether autosomal mo-2 regions experience PAR-like DSB formation, we used maps of ssDNA bound by the strandexchange protein DMC1 (ssDNA sequencing, or SSDS) (9,41,42). Separate studies showed that ANKRD31 is critical for high-level DSB formation in the PAR (30,31). We used our SSDS maps from wholetestis samples of juvenile mice (12 days post partum (30)) and other SSDS maps to test for PAR-like (i.e., ANKRD31-dependent) DSB signal at autosomal mo-2 regions.…”