Abstract
PR domain-containing protein 9 (PRDM9) is a major regulator of the localization of meiotic recombination hotspots in the human and mouse genomes. This role involves its DNA-binding domain, which is composed of a tandem array of zinc fingers, and PRDM9-dependent trimethylation of histone H3 at lysine 4. PRDM9 is a member of the PRDM family of transcription regulators, but unlike other family members, it contains a Krüppel-associated box (KRAB)-related domain that is predicted to be a potential protein interaction domain. Here, we show that truncation of the KRAB domain of mouse PRDM9 leads to loss of PRDM9 function and altered meiotic prophase and gametogenesis. In addition, we identified proteins that interact with the KRAB domain of PRDM9 in yeast two-hybrid assay screens, particularly CXXC1, a member of the COMPASS complex. We also show that CXXC1 interacts with IHO1, an essential component of the meiotic double-strand break (DSB) machinery. As CXXC1 is orthologous to Saccharomyces cerevisiae Spp1 that links DSB sites to the DSB machinery on the chromosome axis, we propose that these molecular interactions involved in the regulation of meiotic DSB formation are conserved in mouse meiosis.
Highlights
Meiotic recombination is initiated by programmed DNA double-strand breaks (DSBs), generated by SPO11 and accessory proteins
The PRDM9 Krüppel-associated box (KRAB) domain is essential for meiosis
To determine its function in the mouse, we introduced a deletion in the KRAB domain by injecting Cas9 mRNA and a single-guide RNA that targets the 5′ site of Prdm9 in C57BL/6(B6) × CBA F1 zygotes (Supplementary Fig. 1b, c)
Summary
Meiotic recombination is initiated by programmed DNA double-strand breaks (DSBs), generated by SPO11 and accessory proteins (de Massy 2013). This KRABrelated domain is present only in PRDM9 and PRDM7 among all PRDM family proteins (Hohenauer and Moore 2012), and its role is unknown We hypothesized that it could be involved in the direct interaction between PRDM9 and a component (s) that enables the recruitment of hotspots to DSB proteins on the chromosome axis. To test this hypothesis, we generated a deletion in Prdm that leads to a truncation of this domain and showed that the PRDM9 KRAB domain is essential for meiosis progression, meiotic DSB repair, and synapsis in both female and male mice. The finding that CXXC1, the mammalian orthologue of S. cerevisiae Spp, interacts with the PRDM9 KRAB domain and with IHO1 leads us to propose a role for PRDM9 KRAB in linking DSB hotspots to the chromosome axis
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