Sex ratio drive in mice: A binding competition between sex-linked genes

Sex chromosomes evolve through dynamic, and often opposing, interplay between sexual selection and sexual conflict, natural selection and intragenomic conflict (1–3). The latter manifests as sex ratio drive, a phenomenon in which a driving locus on a sex chromosome biases the transmission of that chromosome in the germline. When this occurs in the heterogametic sex—XY individuals for example—drive causes biased sex ratios. As the driving locus rises in frequency, producing more of the same sex in a population that already has plenty reduces host fitness. Therefore, selection acting at the level of the host favors any mutation that suppresses the selfish behavior of the driver and recalibrates the sex ratio (1). This means that uncovering sex ratio drive usually requires a perturbation that disrupts the antagonistic equilibrium between driver and suppressor. Nearly three decades ago, a study conducted in Paul Burgoyne’s lab described the phenotypic effects of one such perturbation—a chromosomal mutation that deleted a large interval on the long arm of the mouse Y chromosome (4). It was already known that the mouse Y carried genes with functions in spermiogenesis (postmeiotic sperm development) so it was no surprise that mice with the deletion had a variety of sperm abnormalities. Conway et al. (4) identified a candidate spermiogenesis gene that exists in multiple copies on intact mouse Y chromosomes and is normally expressed in postmeiotic round spermatids. More surprising was the finding that mice with the Y deletion produced litters with female-biased sex ratios. The authors speculated that this effect of Y deletion might be a manifestation of a recently proposed model for sex ratio drive (5), in which a selfish X-linked driver is kept in check by a multicopy suppressor on the Y. So begins the story of Sly (Sycp3-like Y-linked), a gene with more than a hundred copies on the mouse Y chromosome and a starring role in a new study in this issue of PNAS by Arlt et al. (6).

To set the stage, a brief recap of prequels to the current study is in order. First, the initial speculation that a multicopy Y-linked gene might be involved in conflict with a gene or genes on the X chromosome was correct. Deletion or suppression of Sly results in sperm abnormalities, female-biased litters, and upregulation of sex-linked genes in postmeiotic germs cells (7–10). Conversely, deletion or suppression of the Slx/Slxl1 (Sycp3-like X-linked and Slx-like 1) gene family on the X results in sperm abnormalities, male-biased litters, and altered expression of sex-linked and autosomal postmeiotic genes (9, 11). In the nuclei of postmeiotic spermatids, SLY protein colocalizes to the sex chromosomes with epigenetic markers of transcriptional repression and is found in the promoter regions of genes that regulate expression and chromatin remodeling (8, 10). So it seems that Sly enforces Mendel’s law of random segregation by maintaining partial epigenetic repression of Slx/Slxl1 and other sex-linked genes, whereas Slx/Slxl1 biases X chromosome transmission when Sly-mediated repression is disabled.

This competition between X- and Y-linked genes is very much a family feud. The oldest family member, Slxl1, is derived from Synaptonemal complex protein 3 (Sycp3) (11), a single copy autosomal gene essential to the formation of the protein scaffold that encases all chromosomes in meiotic prophase I (12). Slxl1 gave rise to Slx on the X (11) and, as demonstrated by Arlt et al. (6), Sly on the Y (Fig. 1). Prior work from the lab in which the current study was done revealed no evidence for direct interaction between SLY and SLX/SLXL1 proteins and instead identified spindlin proteins, SPIN1 (single copy autosomal) and SSTY1 and SSTY2 (multicopy on the Y), as the probable molecular substrates of competition between SLY and SLX/SLXL1 (11) (see also ref. 13). Given that spindlins regulate chromatin architecture, this seemed consistent with the effects of Sly or Slx/Slxl1 perturbation on gene expression.

Fig. 1.

Evolutionary steps in the arms race between the X and Y chromosomes in Mus musculus. Gene copy numbers for Sstx and Ssty1/2 are from ref. 14.

Arlt et al. (6) begin by digging deeper into the mechanistic basis of X-Y competition. Are SPIN1, SSTY1, and SSTY2 all competitively bound by SLY and SLX/SLXL1? Are SLX and SLXL1 interchangeable in this competition? Is the same true of the two isoforms of SLY, SLY1, and SLY2? The answer to all these questions is no. Using a yeast two-hybrid assay, the authors demonstrate that SLXL1 and SLY1 and SLY2 interact with SPIN1 whereas SLX and SLY2 interact with SSTY2. So teams X and Y are each represented by two players with similar but non-interchangeable roles.

Arlt et al. (6) go on to test whether competition between SLX/SLXL1 and SLY1/SLY2 is dose-dependent (Fig. 2). This is a critical question since the amplification of these genes on their respective sex chromosomes certainly seems like a signature of antagonistic coevolution where an increase in copy number on one sex chromosome enhances spindlin-binding capacity, and therefore drives competitor copy number increase on the other sex chromosome. Support for this idea would require that incremental suppression of team Y members should result in incremental increase in spindlin-binding by relevant team X members and vice versa. Ingenious modification of a yeast three-hybrid assay demonstrates that competition between SLXL1 and both SLY1 and SLY2 for SPIN1 binding is indeed dose-dependent. The evidence for dose-dependent competition between SLX and SLY2 for SSTY2 is less strong. While complete suppression of SLY2 significantly increases SLX–SSTY2 binding, partial suppression does not. This result is open to interpretation. Perhaps Sly amplification is largely driven by competition with SLXL1 for SPIN1. Or perhaps SSTY2 is predisposed to preferentially bind SLY2. After all, both Y-linked gene families benefit when X chromosome drive is suppressed. We return to this possible plot twist below.

Fig. 2.

Tug-of-war between sex chromosomes over spindlin protein binding. (A) SLXL1 competes with SLY1/2 to bind with SPIN1. (B) SLX and SLY2 compete to bind with SSTY2.

Prior work yielded evidence suggestive of positive selection on Sly and Slx coding sequence (11). Here, having identified the specific coding regions in Sly, Slx/Slxl1, Spin1, and Ssty2 that are necessary and sufficient for interactions between these proteins, Arlt et al. (6) ask whether positively selected substitutions are found in the relevant intervals. Slx and Sly2, competitors for Ssty2 binding, both show evidence of strong positive selection at a small number of sites in and near focal intervals. Intriguingly, so does Ssty2.

These results raise some interesting questions. First, at what level is selection acting on these genes? Selection will favor an allele that increases its own transmission to the next generation, regardless of any subsequent cost to its host. Left to its own devises, a driving sex-linked allele can theoretically drive itself to extinction (along with the population it invades) by eliminating the opposite sex (1, 15). However, in a sex-biased population, a mutation that increases production of the rarer sex by suppressing drive will be strongly favored by selection acting at the level of the host (15). One interpretation of the coamplification of Slxl1/Slx and Sly is that the Slxl1/Slx gene families are the selfish drivers and Sly is the suppressor that maintains sex ratio parity. In this scenario, evidence for positive selection on Sly would reflect selection on the host. Arlt et al. (6) provide evidence for concurrent amplification of Slxl1 and origin of Sly in a Mus musculus relative, M. caroli, a pattern consistent with Slxl1 as the original driver. But what is Sly really up to? While the fitness of autosomal suppressors of sex ratio drive is aligned with that of the host, a suppressor on the other sex chromosome enjoys higher fitness when it drives on its own behalf. A landmark study on the Slxl1/Slx/Sly system demonstrated that suppression of either Slxl1/Slx or Sly expression resulted in abnormal sperm morphology and biased sex ratios, whereas concurrent suppression of both Slxl1/Slx and Sly effectively restored a balanced sex ratio and normal sperm morphology (9). These results are certainly suggestive of a pair of sex-linked drivers that are individually damaging and collectively nonessential to host fitness. Second, the other Y-linked protagonist in this story, Ssty2, also has an amplified paralog on the X (Sstx) and is thought to have a history of drive/drive suppression that predates the amplification of Slxl1 and origin of Sly (16). Whether the high copy number of the Ssty1/Ssty2 gene family on the mouse Y reflects an earlier conflict with Sstx or involvement in the more recent conflict between Slxl1/Slx and Sly is currently unclear. Future study of the sum of Ssty interaction partners and the effects of Ssty ablation on sex ratio would help to address these questions. As noted above, physical linkage on the nonrecombining Y means that a transmission advantage gained by Sly benefits Ssty and vice versa. It is tempting to speculate that the positively selected sites in Sly2 and Ssty2 enhance the specificity of interaction between these proteins, a possibility that could be tested in vitro with site-directed mutagenesis.

Finally, the Arlt et al. (6) study provokes broader questions about the drivers of gene transposition, duplication, and amplification on the sex chromosomes in general and the mammalian Y chromosome in particular.

Finally, the Arlt et al. (6) study provokes broader questions about the drivers of gene transposition, duplication, and amplification on the sex chromosomes in general and the mammalian Y chromosome in particular. All Y chromosomes studied to date harbor lineage- and in some cases species-specific genes that have arrived from elsewhere in the genome and proliferated on the Y in recent evolutionary time (14, 17). The lack of recombination, infestation with transposable elements, and reduced effective population sizes of mammalian Y chromosomes make the survival of these added genes a bit of a mystery. To what extent does sex ratio drive contribute to gene survival on the Y? Are some genes functionally predisposed to generate conflict between the sex chromosomes? It is certainly intriguing that the ancestry of Slxl1/Slx/Sly traces to Sycp3, a meiotic gene whose ablation in female mice leads to missegregation of homologous chromosomes (18). Like any good study, the results reported by Arlt et al. (6) answer some questions and raise even more. We will just have to wait for the sequel to see how this particular drama in the male germline unfolds. The complexity of conflict between multicopy genes on the mouse sex chromosomes is certainly suggestive of an evolutionary arms race that is both ancient and ongoing.