Research ArticleGENETICS

Sexual dimorphism in the meiotic requirement for PRDM9: A mammalian evolutionary safeguard

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Science Advances  23 Oct 2020:
Vol. 6, no. 43, eabb6606
DOI: 10.1126/sciadv.abb6606
  • Fig. 1 Prdm9EP alters the Prdm9-dependent H3K4me3 and meiotic DSB landscape in spermatocytes.

    (A and B) Genome browser snapshots of H3K4me3 and DMC1 ChIP-seq peaks in wild-type and Prdm9EP/EP (Glu360Pro mutation) spermatocytes. (A) One strong and one weaker PRDM9-dependent hotspot are shown, together with one PRDM9-independent H3K4me3 peak. (B) Several PRDM9-dependent and PRDM9-independent H3K4me3 peaks are shown. Note the shift in DMC1 peaks to PRDM9-independent H3K4me3 peaks in Prdm9EP/EP spermatocytes. (C) Venn diagrams showing the number of detectable PRDM9-dependent H3K4me3 peaks in wild-type (B6) and Prdm9EP/EP spermatocytes. (D) Venn diagram directly comparing the number of detectable PRDM9-dependent H3K4me3 peaks in wild-type and Prdm9EP/EP spermatocytes. (E) MA plot comparing Prdm9EP/EP versus wild-type signal at PRDM9-dependent H3K4me3 peaks that were detectable in Prdm9EP/EP spermatocytes (n = 3703). (F) Aggregation plot showing normalized average signal intensity [reads per million (RPM)] at known PRDM9-dependent H3K4me3 ChIP-seq peaks (n = 18,838) in wild-type and Prdm9EP/EP spermatocytes. (G) Aggregation plot showing normalized average signal intensity (reads per million, RPM) at common nonhotspot H3K4me3 ChIP-seq peaks (n = 79,043) in wild-type and Prdm9EP/EP spermatocytes. (H) Distribution of DMC1 peaks in wild-type and Prdm9EP/EP spermatocytes. (I) Venn diagram comparing the number of PRDM9-dependent DMC1 peaks in wild-type and Prdm9EP/EP spermatocytes. (J and K) MA plots comparing Prdm9EP/EP versus wild-type DMC1 signal at (J) common PRDM9-dependent DMC1 peaks (n = 4724) and (K) common PRDM9-independent DMC1 peaks (n = 378). Black points represent autosomal peaks, and purple points represent peaks on the X chromosome. EP, Glu360Pro change in amino acid sequence.

  • Fig. 2 Females homozygous for Prdm9EP are fertile.

    (A) Periodic acid–Schiff (PAS)–stained sections from 3-week postpartum ovaries in wild-type and Prdm9EP/EP mice in the B6 genetic background. Arrows show primary follicles. Scale bars, 100 μm. (B) Oocytes per ovary in wild-type, heterozygous, and Prdm9EP/EP females (error bars, SEM). P values were not calculated as N = 2 is not sufficient for reaching statistical significance. (C) Litter sizes in wild-type, Prdm9−/−, and Prdm9EP/EP female mice (error bars, SEM). P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. (D) Pups produced by Prdm9EP/EP female mice. (E) Coimmunolabeling detection of RNF212 foci (red, top row), MLH1 (red, bottom row), and SYCP3 (blue) in pachytene oocyte chromatin spreads in wild-type and mutant females. White arrows highlight unsynapsed regions of chromosomes. Scale bars, 10 μm. (F) Violin plot with dots showing numbers of MLH1 foci per meiotic oocyte (error bars, SEM), in wild-type, Prdm9−/−, and Prdm9EP/EP mice. P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. (G) Diagram showing proportions of PRDM9-dependent and PRDM9-independent crossovers in progeny of B6WSBF2.Prdm9EP/EP females (n = 94). Photo credit: Natalie R. Powers and Tanmoy Bhattacharyya, The Jackson Laboratory.

  • Fig. 3 CAST/EiJ Prdm9-null females are fertile.

    (A) Immunofluorescence staining of histology sections from 3-week postpartum ovaries in wild-type and Prdm9−/− females of different genetic backgrounds. DDX4 (red, also known as MVH) marks oocytes. DNA is stained with 4′,6-diamidino-2-phenylindole (DAPI) (blue). Arrowheads indicate primordial follicles. Scale bars, 100 μm. Note the oocyte depletion in Prdm9−/− mice in the B6 and C3H genetic backgrounds, in contrast to survival of oocytes in wild-type and Prdm9−/− mice in the CAST genetic background. (B) Pups produced by CAST.Prdm9−/− female mice. (C) Reproductive productivity of wild-type and Prdm9−/− females in different genetic backgrounds. P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. NS, not significant. (D) Coimmunolabeling detection of MLH1 foci (red) in pachytene oocyte chromatin spreads, also labeled with antibody against SYCP3 (blue) in wild-type and Prdm9−/− CAST females. Scale bars, 10 μm. (E) Violin plot with dots showing numbers of MLH1 foci per oocyte (error bars, SEM). The genotypes of mice tested are indicated below the graphs. P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. (F) Scheme of construction of B6CASTF2.Prdm9−/− female cohorts used for SNP array genotyping [Giga Mouse Universal Genotyping Array (GigaMUGA)]. Seventy-five F2 females were generated by crossing B6CASTF1.Prdm9−/− and B6CASTF1.Prdm9+/− females with B6CASTF1.Prdm9+/− males. Eight-week-old B6CASTF2.Prdm9−/− females were phenotyped for oocyte quantity and genotyped. (G) Quantitative trait locus (QTL) mapping, with the total number of ovarian follicles (refer to Materials and Methods for details) per B6CASTF2.Prdm9−/− female as the phenotype. QTL (pink) reached significance on chromosome 5, with a peak at ~100.4 Mb (1.5 LOD drop: 72.9 to 127.65 Mb, mm10). (H) Genotype effects of the chromosome 5 QTL. CAST alleles affect oocyte number positively, as highlighted by CC (violet) and BC (orange) lines, relative to B6 homozygotes (BB, green). CC, homozygous CAST; BB, homozygous B6; BC, heterozygous CAST/B6. Photo credit: Catherine Brunton and Tanmoy Bhattacharyya, The Jackson Laboratory.

  • Fig. 4 Chk2 ablation rescues meiosis in Prdm9-null females in the B6 genetic background.

    (A and C) Coimmunolabeling detection of MLH3 foci (red, A), MLH1 (red, C), and SYCP3 (blue, both A and C) in pachytene oocyte chromatin spreads from wild-type and mutant females. Yellow arrowheads highlight unsynapsed regions of chromosomes. Scale bars, 10 μm. (B and D) Violin plots with dots showing numbers of MLH3 (B) and MLH1 (D) foci per meiotic oocyte (error bars, SEM). The genotypes of mice tested are indicated below the graphs. P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. (E and F) Chromosome configuration in meiosis MI analyzed by chromosome spreads from different mutant and control oocytes, pictured in (E) and quantified in (F). In (E), DNA (blue) and kinetochores (red) were detected by DAPI and CREST antiserum, respectively. Scale bars,10 μm. Multiple univalents in a B6.Prdm9−/− oocyte are indicated with white arrowheads. The examples of oocytes from B6 and B6.Prdm9−/−Chk2−/− mice reveal all chromosomes organized in bivalents. P values were calculated using χ2 test.

  • Fig. 5 Chk2 ablation rescues fertility in Prdm9-null females in the B6 genetic background.

    (A) PAS-stained histological sections of 3-week postpartum ovaries from wild-type, single-, and double-mutant females in the B6 genetic background. Scale bars, 200 μm. (B) Pups produced by B6.Prdm9−/−Chk2−/− female mice. (C) Oocyte quantification in mutant and wild-type animals. (D) Litter sizes in mutant and control females. P values were calculated using the Mann-Whitney U test with Bonferroni correction for multiple testing. Photo credit: Tanmoy Bhattacharyya, The Jackson Laboratory.

Supplementary Materials

  • Supplementary Materials

    Sexual dimorphism in the meiotic requirement for PRDM9: A mammalian evolutionary safeguard

    Natalie R. Powers, Beth L. Dumont, Chihiro Emori, Raman Akinyanju Lawal, Catherine Brunton, Kenneth Paigen, Mary Ann Handel, Ewelina Bolcun-Filas, Petko M. Petkov, Tanmoy Bhattacharyya

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