Re-engineering the zinc fingers of PRDM9 reverses hybrid sterility in mice

Benjamin Davies,Julie G Hussin,Chris Preece,Florencia Pratto,Daniel Biggs,Kevin Brick,Catherine M Green,Emmanuelle Bitoun,R Daniel Camerini-Otero,Gang Zhang,Rebeca Diaz,Ran Li,Nicolas Altemose,Simon R Myers,Daniela Moralli,Edouard Hatton,Anjali Gupta Hinch,Peter Donnelly

doi:10.1038/nature16931

Abstract

The DNA-binding protein PRDM9 directs positioning of the double-strand breaks (DSBs) that initiate meiotic recombination in mice and humans. Prdm9 is the only mammalian speciation gene yet identified and is responsible for sterility phenotypes in male hybrids of certain mouse subspecies. To investigate PRDM9 binding and its role in fertility and meiotic recombination, we humanized the DNA-binding domain of PRDM9 in C57BL/6 mice. This change repositions DSB hotspots and completely restores fertility in male hybrids. Here we show that alteration of one Prdm9 allele impacts the behaviour of DSBs controlled by the other allele at chromosome-wide scales. These effects correlate strongly with the degree to which each PRDM9 variant binds both homologues at the DSB sites it controls. Furthermore, higher genome-wide levels of such 'symmetric' PRDM9 binding associate with increasing fertility measures, and comparisons of individual hotspots suggest binding symmetry plays a downstream role in the recombination process. These findings reveal that subspecies-specific degradation of PRDM9 binding sites by meiotic drive, which steadily increases asymmetric PRDM9 binding, has impacts beyond simply changing hotspot positions, and strongly support a direct involvement in hybrid infertility. Because such meiotic drive occurs across mammals, PRDM9 may play a wider, yet transient, role in the early stages of speciation.

Highlights

The DNA-binding protein PRDM9 directs positioning of the double strand breaks (DSBs) initiating meiotic recombination, in mice and humans
Humanization of the Prdm[9] zinc finger (ZF)-array in B6 inbred mice had no effect on fertility (Extended Data Fig. 2) and cytogenetic comparisons revealed no significant impact on zygotene DSB counts (DMC1 immunoreactivity, Extended Data Fig. 2b), crossover counts (MLH1 foci, Extended Data Fig. 2c), normal sex body formation or quantitative measures of fertility and successful synapsis
We found that most DMC1 signal (71.8%) in (PWD×B6)F1PWD/B6 or (B6×PWD)F1B6/PWD hybrids occurs within asymmetric DSB hotspots (Fig. 2a, Extended Data Fig. 6)

Summary

Discussion

One mammalian speciation gene, Prdm[9], has so far been identified. Humanizing the ZF-array of Prdm[9] redirects binding, thereby entirely reprogramming recombination hotspots and in doing so reverses the hybrid infertility between musculus and domesticus subspecies. It has been suggested that synaptonemal complex proteins are loaded at some DSB sites and synapsis begins to spread[7,8] Extending this model, to incorporate the property that asymmetrically bound sites are less favourable for homology search, would parsimoniously predict each symmetry-related phenomenon we observed: DSBs at asymmetric hotspots would repair more slowly, elevating their DMC1 signal, and chromosomes with fewer symmetric hotspots overall would show delayed DSB repair and higher asynapsis rates, causing subfertility or sterility in animals with low symmetric binding. Hotspots might massively increase search efficiency by directing homology search to PRDM9 binding sites

Methods

Findings

Analysis code availability and source data