ATM and PRDM9 Regulate SPO11-bound Recombination Intermediates During Meiosis

Abstract

Meiotic recombination is initiated by genome-wide SPO11-induced double-strand breaks (DSBs) that are processed by MRE11-mediated release of SPO11. The DSB is then resected and loaded with DMC1/RAD51 filaments that invade homologous chromosome templates. In most mammals, DSB locations ("hotspots") are determined by the DNA sequence specificity of PRDM9. Here, we demonstrate the first direct detection of meiotic DSBs and resection in vertebrates by performing END-seq on mouse spermatocytes using low sample input. We find that DMC1 limits both the minimum and maximum lengths of resected DNA, whereas 53BP1, BRCA1 and EXO1 play surprisingly minimal roles in meiotic resection. Through enzymatic modifications to the END-seq protocol that mimic the in vivo processing of SPO11, we identify a novel meiotic recombination intermediate ("SPO11-RI") present at all hotspots. The SPO11-bound intermediate is dependent on PRDM9 and caps the 3 resected end during engagement with the homologous template. We propose that SPO11-RI is generated because chromatin-bound PRDM9 asymmetrically blocks MRE11 from releasing SPO11. In Atm-/- spermatocytes, SPO11-RI is reduced while unresected DNA-bound SPO11 accumulate because of defective MRE11 initiation. Thus in addition to their global roles in governing SPO11 breakage, ATM and PRDM9 are critical local regulators of mammalian SPO11 processing.