Interrogating the Functions of PRDM9 Domains in Meiosis

Abstract

Homologous recombination is required for proper segregation of homologous chromosomes during meiosis. It occurs predominantly at recombination hotspots that are defined by the DNA binding specificity of the PRDM9 protein. PRDM9 contains three conserved domains typically involved in regulation of transcription; yet, the role of PRDM9 in gene expression control is not clear. Here, we analyze the germline transcriptome of Prdm9−/− male mice in comparison to Prdm9+/+ males and find no apparent differences in the mRNA and miRNA profiles. We further explore the role of PRDM9 in meiosis by analyzing the effect of the KRAB, SSXRD, and post-SET zinc finger deletions in a cell culture expression system and the KRAB domain deletion in mice. We found that although the post-SET zinc finger and the KRAB domains are not essential for the methyltransferase activity of PRDM9 in cell culture, the KRAB domain mutant mice show only residual PRDM9 methyltransferase activity and undergo meiotic arrest. In aggregate, our data indicate that domains typically involved in regulation of gene expression do not serve that role in PRDM9, but are likely involved in setting the proper chromatin environment for initiation and completion of homologous recombination.

ZCWPW1 is recruited to recombination hotspots by PRDM9, and is essential for meiotic double strand break repair

“…This implies that ZCWPW1 is most often lost, whenever mutations occur in the SET domain of PRDM9. Interestingly, PRDM9-containing species with non-mutant SET domains are near-identical to those also possessing SSXRD domains, and SSXRD has been reported as essential for PRDM9's H3K4me3 methyltransferase activity at hotspots ex vivo (Thibault-Sennett et al, 2018). However, other domains of PRDM9 (KRAB, and the zinc finger array) have been lost across multiple species, and it is hypothesized PRDM9 does not position recombination in these species (Baker et al, 2017).…”

HELLS and PRDM9 form a Pioneer Complex to Open Chromatin at Meiotic Recombination Hotspots

“…Expression of PRDM9 ex vivo recapitulates aspects of hotspot activation including allelespecific H3K4me3 deposition (Altemose et al, 2017;Baker et al, 2015b;Thibault-Sennett et al, 2018). To develop a model for temporal molecular characterization of hotspot activation, we choose a human cell line that expresses HELLS and created a stably-integrated FLAG-PRDM9 C allele under inducible control of the tetracycline promoter (HEK293-P9 C , Fig.…”

Uncoupling of transcriptomic and cytological differentiation in mouse spermatocytes with impaired meiosis

“…This gene encodes PRDM9 (PR/SET Domain 9), a zinc-finger DNA-binding protein with histone methyltransferase activity that is expressed in early meiotic prophase, during leptonema and zygonema (Sun et al , 2015), and is required for activation of recombination (Parvanov et al , 2010). Recent work has demonstrated that, despite its deposition of classically gene-activating marks, PRDM9 does not directly regulate gene expression (Thibault-Sennett et al , 2018). Mice bearing inactivating mutations of Prdm9 (herein designated as Prdm9 –/– ) are infertile and meiosis is arrested at early to midprophase, with the failure of reciprocal recombination and absence of cytologically normal germ cells past the zygotene substage.…”