Genic and chromosomal components of Prdm9-driven hybrid male sterility in mice (Mus musculus)

Abstract

Hybrid sterility contributes to speciation by preventing gene flow between related taxa. Prdm9, the first and only hybrid male sterility gene known in vertebrates, predetermines the sites of recombination between homologous chromosomes and their synapsis in early meiotic prophase. The asymmetric binding of PRDM9 to heterosubspecific homologs of Mus musculus musculus × Mus musculus domesticus F1 hybrids and increase of PRDM9-independent DNA double-strand break hotspots results indificult- to- repair double-strand breaks, incomplete synapsis of homologous chromosomes, and meiotic arrest at the first meiotic prophase. Here, we show that Prdm9 behaves as a major hybrid male sterility gene in mice outside the Mus musculus musculus × Mus musculus domesticus F1 hybrids, in the genomes composed of Mus musculus castaneus and Mus musculus musculus chromosomes segregating on the Mus musculus domesticus background. The Prdm9cst/dom2 (castaneus/domesticus) allelic combination secures meiotic synapsis, testes weight, and sperm count within physiological limits, while the Prdm9msc1/dom2 (musculus/domesticus) males show a range of fertility impairment. Out of 5 quantitative trait loci contributing to the Prdm9msc1/dom2-related infertility, 4 control either meiotic synapsis or fertility phenotypes and 1 controls both, synapsis, and fertility. Whole-genome genotyping of individual chromosomes showed preferential involvement of nonrecombinant musculus chromosomes in asynapsis in accordance with the chromosomal character of hybrid male sterility. Moreover, we show that the overall asynapsis rate can be estimated solely from the genotype of individual males by scoring the effect of nonrecombinant musculus chromosomes. Prdm9-controlled hybrid male sterility represents an example of genetic architecture of hybrid male sterility consisting of genic and chromosomal components.

Keywords: HORMAD2; SYCP3; homologous synapsis; meiosis; spermatogenesis; synaptonemal complex.

Fig. 1.

Prdm9 gene and Hstx2 locus control fertility of (musculus^PWD × castaneus^CAST) × domesticus^B6 TC1 males. a) The Prdm9^cst/dom2 males are all fertile, while the Prdm9^msc1/dom2 allelic combination determines variable fertility phenotypes ranging from complete sterility to fertility. This is in contrast to (musculus^PWD × domesticus^B6)F1 hybrids where the same Prdm9^msc1/dom2 allelic combination causes complete male sterility (insert). b) The Hstx2^CAST allele moderately reduces weight of the testes in fertile Prdm9^cst/dom2 hybrid males, but c) it does not significantly change the number of sperm in the epididymis. Prdm9 c, d, and m refers to cst, dom2, and msc1 alleles of castaneus, domesticus and musculus origin. Hstx2 c and m refers to cst and msc1 alleles of castaneus and musculus origin.

Fig. 2.

Standard interval QTL mapping of loci controlling fertility phenotypes in Prdm9^msc1/dom2 TC1 hybrids. a) Testes weight and b) sperm count as fertility phenotypes. Genome-wide significance thresholds (α = 0.05 and α = 0.10) are indicated by the red and blue dashed line, respectively. c) The effect of individual QTLs on the testes weight and the combined effect of twQTLs grouped according CAST (C) or PWD (P) in the order of alleles of twQTL-4, twQTL-18, and twQTL-X loci. The CAST allele of twQTL-X reduces weight of the testes in contrast to the X-linked Hstx2^CAST in the whole set of TC1 males (see Fig. 1c). d) The scQTLs on Chr 4 and 18 display recessive-to-additive epistatic interaction.

Fig. 3.

Control of meiotic chromosome synapsis and fertility phenotypes by Prdm9 and Hstx2 genes. a) The subspecific origin of TC1 autosomes is shown on Chr 17 as an example. The probability of homologous synapsis failure mainly depends on the subspecific origin of autosomes and Prdm9 allelic combinations, symbols +, ±, and − refer to high, medium, and low/null probability of asynapsis, respectively. Other factors, such as asyQTLs and MSUC, are not considered here. b) Pachytene spread showing synaptonemal complexes in a fully synapsed cell (upper panel) and a cell with 3 asynapsed pairs of autosomes (lower panel). X–Y chromosomes and asynapsed autosomes are decorated with anti-HORMAD2 antibody (yellow–green). Note weak HORMAD2 staining on the XY chromosomes indicating incomplete MSCI. CEN labels centromeric heterochromatin, and DAPI labels nuclear DNA. c) Relation between asynapsis and testes weight and d) asynapsis and sperm count in Prdm9^cst/dom2 (orange) and Prdm9^msc1/dom2 (blue) TC1 males. e) The Hstx2^CAST allele or a closely linked gene increases asynapsis rate in sterile and fertile TC1 hybrids.

Fig. 4.

Meiotic asynapsis as a quantitative trait in Prdm9^msc1/dom2 TC1 hybrids. a) Standard interval QTL mapping of loci controlling meiotic chromosome asynapsis. The genome-wide significance threshold (α = 0.05 and α  =  0.10) is indicated by the red and blue dashed lines, respectively. b) The asyQTL allelic combinations show additive epistatic interaction between loci on Chrs 3, 18, and X. The average asynapsis rates are shown for subsets of males carrying the CAST allele in all 3, 2, 1, or none asyQTLs (C3_P0, C2_P1, C1_P2, and C_P3). Vertical bars designate 5–95 percentile, and horizontal bars represent medians.

Fig. 5.

Relative asynapsis rate of 3 most sensitive autosomes in Prdm9^msc1/dom2 TC1 males (see Bhattacharyya et al. 2013; Kauppi et al. 2013; Gregorova et al. 2018). a, b) The average asynapsis rate was estimated by visualization of Chrs 17, 18, and 19 by DNA FISH. a) Fully synapsed Chr 17 is decorated by a Chr 17-specific DNA-painting probe. b) Asynapsed Chr 19 often associates with Chrs X and Y. c) The effect of the CAST sequence on the asynapsis rate of Chrs 17, 18, and 19 in individual TC1 males. The subspecies origin of individual chromosomes coming from a (PWD × CAST)F1 female is either nonrecombinant PWD (P), nonrecombinant CAST (C), recombinant PWD–CAST (P–C, centromere to telomere) or recombinant CAST–PWD (C–P). The length of these chromosomal intervals is given in Mb (genome assembly: GRCm39).

Fig. 6.

Spearman correlation between a) predicted and observed asynapsis (Spearman 0.67, RMSE = 0.159) and b) between predicted, QTL-adjusted, and observed asynapsis rate (Spearman 0.77, RMSE = 0.126). The QTL-adjusted, predicted asynapsis rate is significantly closer to the observed asynapsis (Mann–Whitney P = 0.012).

Fig. 7.

Distribution of testes weights of Prdm9^msc1/dom2 TC1 males with contrasting allelic combinations of twQTLs. a) Gray columns represent all TC1 males tested for twQTL-4, twQTL-18, and twQTL-X. The dark blue columns show testes weights of males with twQTL-4^PWD, twQTL-18^PWD, and twQTL-X^CAST allelic combination (“sterile” allelic combination), while light blue columns refer to twQTL-4^CAST, twQTL-18^CAST, and twQTL-X^PWD males (“fertile” combination). b) The predicted asynapsis rate correlates with testes weight in the “fertile” combination group (blue dots, Spearman r = −0.786, P = 0.0035, 2-tailed), but the correlation is lost in the “sterile” group (orange squares, Spearman r = −0.22, P = 0.34, 2-tailed) indicating the dominance of genic twQTL determinants over chromosomal incompatibility.

Fig. 8.

Schematic representation of the events following PRDM9-driven DNA DSB induction during the first meiotic prophase of Prdm9^msc1/dom2 intersubspecific male hybrids. The scheme is based on the DNA DSB asymmetry hypothesis (Davies et al. 2016; Gregorova et al. 2018). Increasingly unilateral binding of PRDM9 to its nonself homolog (musculus PRDM9 on domesticus chromosome and vice versa) can increase the risk of delay or failure of DSB repair and immediate cell death before meiotic pairing or can result in pachynemas with incomplete synapsis of homologous chromosomes. Failure to synapse can trigger the pachytene checkpoint and induce meiotic silencing of unsynapsed chromatin (MSUC), resulting in transcriptional silencing in cis of genes essential for meiosis, or in trans by failed transcriptional inactivation of sex chromosomes (MSCI).