A complex genetic basis to X-linked hybrid male sterility between two species of house mice

The X chromosome plays a central role in the evolution of reproductive isolation, but few studies have examined the genetic basis of X-linked incompatibilities during the early stages of speciation. We report the results of a large experiment focused on the reciprocal introgression of the X chromosome between two species of house mice, Mus musculus and M. domesticus. Introgression of the M. musculus X chromosome into a wild-derived M. domesticus genetic background produced male-limited sterility, qualitatively consistent with previous experiments using classic inbred strains to represent M. domesticus. The genetic basis of sterility involved a minimum of four X-linked factors. The phenotypic effects of major sterility QTL were largely additive and resulted in complete sterility when combined. No sterility factors were uncovered on the M. domesticus X chromosome. Overall, these results revealed a complex and asymmetric genetic basis to X-linked hybrid male sterility during the early stages of speciation in mice. Combined with data from previous studies, we identify one relatively narrow interval on the M. musculus X chromosome involved in hybrid male sterility. Only a handful of spermatogenic genes are within this region, including one of the most rapidly evolving genes on the mouse X chromosome.     

Genetic dissection of a key reproductive barrier between nascent species of house mice

Reproductive isolation between species is often caused by deleterious interactions among loci in hybrids. Finding the genes involved in these incompatibilities provides insight into the mechanisms of speciation. With recently diverged subspecies, house mice provide a powerful system for understanding the genetics of reproductive isolation early in the speciation process. Although previous studies have yielded important clues about the genetics of hybrid male sterility in house mice, they have been restricted to F1 sterility or incompatibilities involving the X chromosome. To provide a more complete characterization of this key reproductive barrier, we conducted an F2 intercross between wild-derived inbred strains from two subspecies of house mice, Mus musculus musculus and Mus musculus domesticus. We identified a suite of autosomal and X-linked QTL that underlie measures of hybrid male sterility, including testis weight, sperm density, and sperm morphology. In many cases, the autosomal loci were unique to a specific sterility trait and exhibited an effect only when homozygous, underscoring the importance of examining reproductive barriers beyond the F1 generation. We also found novel two-locus incompatibilities between the M. m. musculus X chromosome and M. m. domesticus autosomal alleles. Our results reveal a complex genetic architecture for hybrid male sterility and suggest a prominent role for reproductive barriers in advanced generations in maintaining subspecies integrity in house mice.     

The contribution of the y chromosome to hybrid male sterility in house mice

Hybrid sterility in the heterogametic sex is a common feature of speciation in animals. In house mice, the contribution of the Mus musculus musculus X chromosome to hybrid male sterility is large. It is not known, however, whether F(1) male sterility is caused by X-Y or X-autosome incompatibilities or a combination of both. We investigated the contribution of the M. musculus domesticus Y chromosome to hybrid male sterility in a cross between wild-derived strains in which males with a M. m. musculus X chromosome and M. m. domesticus Y chromosome are partially sterile, while males from the reciprocal cross are reproductively normal. We used eight X introgression lines to combine different X chromosome genotypes with different Y chromosomes on an F(1) autosomal background, and we measured a suite of male reproductive traits. Reproductive deficits were observed in most F(1) males, regardless of Y chromosome genotype. Nonetheless, we found evidence for a negative interaction between the M. m. domesticus Y and an interval on the M. m. musculus X that resulted in abnormal sperm morphology. Therefore, although F(1) male sterility appears to be caused mainly by X-autosome incompatibilities, X-Y incompatibilities contribute to some aspects of sterility.     

Genetics and evolution of hybrid male sterility in house mice

Comparative genetic mapping provides insights into the evolution of the reproductive barriers that separate closely related species. This approach has been used to document the accumulation of reproductive incompatibilities over time, but has only been applied to a few taxa. House mice offer a powerful system to reconstruct the evolution of reproductive isolation between multiple subspecies pairs. However, studies of the primary reproductive barrier in house mice-hybrid male sterility-have been restricted to a single subspecies pair: Mus musculus musculus and Mus musculus domesticus. To provide a more complete characterization of reproductive isolation in house mice, we conducted an F(2) intercross between wild-derived inbred strains from Mus musculus castaneus and M. m. domesticus. We identified autosomal and X-linked QTL associated with a range of hybrid male sterility phenotypes, including testis weight, sperm density, and sperm morphology. The pseudoautosomal region (PAR) was strongly associated with hybrid sterility phenotypes when heterozygous. We compared QTL found in this cross with QTL identified in a previous F(2) intercross between M. m. musculus and M. m. domesticus and found three shared autosomal QTL. Most QTL were not shared, demonstrating that the genetic basis of hybrid male sterility largely differs between these closely related subspecies pairs. These results lay the groundwork for identifying genes responsible for the early stages of speciation in house mice.     

GENETICS OF STERILITY IN HYBRIDS BETWEEN TWO SUBSPECIES OF DROSOPHILA

Hybrids between D. pseudoobscura bogotana and D. pseudoobscura pseudoobscura are fertile except for males produced in one of the two reciprocal crosses. As there is no premating isolation between these subspecies, nonreciprocal male sterility represents the first step in speciation. Genetic analysis reveals two causes of hybrid F₁ sterility: a maternal effect and incompatibilities between chromosomes within males. The maternal effect appears to play the greatest role in hybrid sterility. The X chromosome has the largest effect on fertility of any chromosome, a ubiquitous result in analyses of hybrid sterility and inviability in Drosophila. This effect is entirely attributable to a region comprising less than 30% of the X chromosome. These results are compared to those from a similar study of D. pseudoobscura-D. persimilis hybrids, an older and more reproductively isolated species pair in the same lineage. Such comparisons may allow one to identify the genetic changes characterizing the early versus late stages of speciation.     

Reduced male fertility is common but highly variable in form and severity in a natural house mouse hybrid zone

Barriers to gene flow between naturally hybridizing taxa reveal the initial stages of speciation. Reduced hybrid fertility is a common feature of reproductive barriers separating recently diverged species. In house mice (Mus musculus), hybrid male sterility has been studied extensively using experimental crosses between subspecies. Here, we present the first detailed picture of hybrid male fertility in the European M. m. domesticus-M. m. musculus hybrid zone. Complete sterility appears rare or absent in natural hybrids but a large proportion of males (~30%) have sperm count or relative testis weight below the range in pure subspecies, and likely suffer reduced fertility. Comparison of a suite of traits related to fertility among subfertile males indicates reduced hybrid fertility in the contact zone is highly variable among individuals and ancestry groups in the type, number, and severity of spermatogenesis defects present. Taken together, these results suggest multiple underlying genetic incompatibilities are segregating in the hybrid zone, which likely contribute to reproductive isolation between subspecies.     

Signatures of reproductive isolation in patterns of single nucleotide diversity across inbred strains of mice

Reproductive isolation is often caused by the disruption of genic interactions that evolve in geographically separate populations. Identifying the genomic regions and genes involved in these interactions, known as "Dobzhansky-Muller incompatibilities," can be challenging but is facilitated by the wealth of genetic markers now available in model systems. In recent years, the complete genome sequence and thousands of single nucleotide polymorphisms (SNPs) from laboratory mice, which are largely genetic hybrids between Mus musculus and M. domesticus, have become available. Here, we use these resources to locate genomic regions that may underlie reproductive isolation between these two species. Using genotypes from 332 SNPs that differ between wild-derived strains of M. musculus and M. domesticus, we identified several physically unlinked SNP pairs that show exceptional gametic disequilibrium across the lab strains. Conspecific alleles were associated in a disproportionate number of these cases, consistent with the action of natural selection against hybrid gene combinations. As predicted by the Dobzhansky-Muller model, this bias was differentially attributable to locus pairs for which one hybrid genotype was missing. We assembled a list of potential Dobzhansky-Muller incompatibilities from locus pairs that showed extreme associations (only three gametic types) among conspecific alleles. Two SNPs in this list map near known hybrid sterility loci on chromosome 17 and the X chromosome, allowing us to nominate partners for disrupted interactions involving these genomic regions for the first time. Together, these results indicate that patterns produced by speciation between M. musculus and M. domesticus are visible in the genomes of lab strains of mice, underscoring the potential of these genetic model organisms for addressing general questions in evolutionary biology.     

Speciation and reduced hybrid female fertility in house mice

In mammals, intrinsic postzygotic isolation has been well studied in males but has been less studied in females, despite the fact that female gametogenesis and pregnancy provide arenas for hybrid sterility or inviability that are absent in males. Here, we asked whether inviability or sterility is observed in female hybrids of Mus musculus domesticus and M. m. musculus, taxa which hybridize in nature and for which male sterility has been well characterized. We looked for parent‐of‐origin growth phenotypes by measuring adult body weights in F1 hybrids. We evaluated hybrid female fertility by crossing F1 females to a tester male and comparing multiple reproductive parameters between intrasubspecific controls and intersubspecific hybrids. Hybrid females showed no evidence of parent‐of‐origin overgrowth or undergrowth, providing no evidence for reduced viability. However, hybrid females had smaller litter sizes, reduced embryo survival, fewer ovulations, and fewer small follicles relative to controls. Significant variation in reproductive parameters was seen among different hybrid genotypes, suggesting that hybrid incompatibilities are polymorphic within subspecies. Differences in reproductive phenotypes in reciprocal genotypes were observed and are consistent with cyto‐nuclear incompatibilities or incompatibilities involving genomic imprinting. These findings highlight the potential importance of reduced hybrid female fertility in the early stages of speciation.     

Genetic studies of two sister species in the Drosophila melanogaster subgroup, D. yakuba and D. santomea

We performed genetic analysis of hybrid sterility and of one morphological difference (sex-comb tooth number) on D. yakuba and D. santomea, the former species widespread in Africa and the latter endemic to the oceanic island of S?o Tomé, on which there is a hybrid zone. The sterility of hybrid males is due to at least three genes on the X chromosome and at least one on the Y, with the cytoplasm and large sections of the autosomes having no effect. F1 hybrid females carrying two X chromosomes from either species are perfectly fertile despite their genetic similarity to completely sterile F1 hybrid males. This implies that the appearance of Haldane's rule in this cross is at least partially due to the faster accumulation of genes causing male than female sterility. The larger effects of the X and Y chromosomes than of the autosomes, however, also suggest that the genes causing male sterility are recessive in hybrids. Some female sterility is also seen in interspecific crosses, but this does not occur between all strains. This is seen in pure-species females inseminated by heterospecific males (probably reflecting incompatibility between the sperm of one species and the female reproductive tract of the other) as well as in inseminated F1 and backcross females, probably reflecting genetically based incompatibilities in hybrids that affect the reproductive system. The latter 'innate' sterility appears to involve deleterious interactions between D. santomea chromosomes and D. yakuba cytoplasm. The difference in male sex-comb tooth number appears to involve fairly large effects of the X chromosome. We discuss the striking evolutionary parallels in the genetic basis of sterility, in the nature of sexual isolation, and in morphological differences between the D. santomea/D. yakuba divergence and two other speciation events in the D. melanogaster subgroup involving island colonization.     

Genetic basis of hybrid male sterility among three closely related species of Drosophila

The genetic basis of hybrid male sterility among three closely related species, Drosophila bipectinata, D. parabipectinata and D. malerkotliana has been investigated by using backcross analysis methods. The role of Y chromosome, major hybrid sterility (MHS) genes (genetic factors) and cytoplasm (non-genetic factor) have been studied in the hybrids of these three species. In the species pair, bipectinata--parabipectinata, Y chromosome introgression of parabipectinata in the genomic background of bipectinata and the reciprocal Y chromosome introgression were unsuccessful as all males in second backcross generation were sterile. Neither MHS genes nor cytoplasm was found important for sterility. This suggests the involvement of X-Y, X-autosomes or polygenic interactions in hybrid male sterility. In bipectinata--malerkotliana and parabipectinata--malerkotliana species pairs, Y chromosome substitution in reciprocal crosses did not affect male fertility. Backcross analyses also show no involvement of MHS genes or cytoplasm in hybrid male sterility in these two species pairs. Therefore, X- autosome interaction or polygenic interaction is supposed to be involved in hybrid male sterility in these two species pairs. These findings also provide evidence that even in closely related species, genetic interactions underlying hybrid male sterility may vary.     

Postzygotic isolation between the two European subspecies of the house mouse: estimates from fertility patterns in wild and laboratory-bred hybrids

We assessed the fertility (reproductive success, litter size, testis weight, spermatocyte-to-spermatid ratio) of F₁s and backcrosses between different wild-derived outbred and inbred strains of two mouse subspecies, Mus musculus domesticus and M. m. musculus . A significant proportion of the F₁ females between the outbred crosses did not reproduce, suggesting that female infertility was present. As the spermatocyte-to-spermatid ratio was correlated with testis weight, the latter was used to attribute a sterile vs. fertile phenotype to all males. Segregation proportions in the backcrosses of F₁ females yielded 11 (inbred) to 17% (outbred) sterile males, suggesting the contribution of two to three major genetic factors to hybrid male sterility. Only one direction of cross between the inbred strains produced sterile F₁ males, indicating that one factor was borne by the musculus X-chromosome. No such differences were observed between reciprocal crosses in the outbred strains. The involvement of the X chromosome in male sterility thus could not be assessed, but its contribution appears likely given the limited introgression of X-linked markers through the hybrid zone between the subspecies. However, we observed no sterile phenotypes in wild males from the hybrid zone, although testis weight tended to decrease in the centre of the transect.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 84 , 379–393.     

The Genetic Basis of Haldane''s Rule and the Nature of Asymmetric Hybrid Male Sterility among Drosophila Simulans, Drosophila Mauritiana and Drosophila Sechellia

Haldane's rule (i.e., the preferential hybrid sterility and inviability of heterogametic sex) has been known for 70 years, but its genetic basis, which is crucial to the understanding of the process of species formation, remains unclear. In the present study, we have investigated the genetic basis of hybrid male sterility using Drosophila simulans, Drosophila mauritiana and Drosophila sechellia. An introgression of D. sechellia Y chromosome into a fairly homogenous background of D. simulans did not show any effect of the introgressed Y on male sterility. The substitution of D. simulans Y chromosome into D. sechellia, and both reciprocal Y chromosome substitutions between D. simulans and D. mauritiana were unsuccessful. Introgressions of cytoplasm between D. simulans and D. mauritiana (or D. sechellia) also did not have any effect on hybrid male sterility. These results rule out the X-Y interaction hypothesis as a general explanation of Haldane's rule in this species group and indicate an involvement of an X-autosome interaction. Models of symmetrical and asymmetrical X-autosome interaction have been developed which explain the Y chromosome substitution results and suggest that evolution of interactions between different genetic elements in the early stages of speciation is more likely to be of an asymmetrical nature. The model of asymmetrical X-autosome interaction also predicts that different sets of interacting genes may be involved in different pairs of related species and can account for the observation that hybrid male sterility in many partially isolated species is often nonreciprocal or unidirectional.     

Genetic dissection of hybrid incompatibilities between Drosophila simulans and D. mauritiana. I. Differential accumulation of hybrid male sterility effects on the X and autosomes

The genetic basis of hybrid incompatibility in crosses between Drosophila mauritiana and D. simulans was investigated to gain insight into the evolutionary mechanisms of speciation. In this study, segments of the D. mauritiana third chromosome were introgressed into a D. simulans genetic background and tested as homozygotes for viability, male fertility, and female fertility. The entire third chromosome was covered with partially overlapping segments. Many segments were male sterile, while none were female sterile or lethal, confirming previous reports of the rapid evolution of hybrid male sterility (HMS). A statistical model was developed to quantify the HMS accumulation. In comparison with previous work on the X chromosome, we estimate that the X has approximately 2.5 times the density of HMS factors as the autosomes. We also estimate that the whole genome contains approximately 15 HMS "equivalents"-i.e., 15 times the minimum number of incompatibility factors necessary to cause complete sterility. Although some caveats for the quantitative estimate of a 2.5-fold density difference are described, this study supports the notion that the X chromosome plays a special role in the evolution of reproductive isolation. Possible mechanisms of a "large X" effect include selective fixation of new mutations that are recessive or partially recessive and the evolution of sex-ratio distortion systems.     

Differential patterns of introgression across the X chromosome in a hybrid zone between two species of house mice

A complete understanding of the speciation process requires the identification of genomic regions and genes that confer reproductive barriers between species. Empirical and theoretical research has revealed two important patterns in the evolution of reproductive isolation in animals: isolation typically arises as a result of disrupted epistatic interactions between multiple loci and these disruptions map disproportionately to the X chromosome. These patterns suggest that a targeted examination of natural gene flow between closely related species at X-linked markers with known positions would provide insight into the genetic basis of speciation. We take advantage of the existence of genomic data and a well-documented European zone of hybridization between two species of house mice, Mus domesticus and M. musculus, to conduct such a survey. We evaluate patterns of introgression across the hybrid zone for 13 diagnostic X-linked loci with known chromosomal positions using a maximum likelihood model. Interlocus comparisons clearly identify one locus with reduced introgression across the center of the hybrid zone, pinpointing a candidate region for reproductive isolation. Results also reveal one locus with high frequencies of M. domesticus alleles in populations on the M. musculus side of the zone, suggesting the possibility that positive selection may act to drive the spread of alleles from one species on to the genomic background of the other species. Finally, cline width and cline center are strongly positively correlated across the X chromosome, indicating that gene flow of the X chromosome may be asymmetrical. This study highlights the utility of natural populations of hybrids for mapping speciation genes and suggests that the middle of the X chromosome may be important for reproductive isolation between species of house mice.     

A simple genetic incompatibility causes hybrid male sterility in mimulus

Much evidence has shown that postzygotic reproductive isolation (hybrid inviability or sterility) evolves by the accumulation of interlocus incompatibilities between diverging populations. Although in theory only a single pair of incompatible loci is needed to isolate species, empirical work in Drosophila has revealed that hybrid fertility problems often are highly polygenic and complex. In this article we investigate the genetic basis of hybrid sterility between two closely related species of monkeyflower, Mimulus guttatus and M. nasutus. In striking contrast to Drosophila systems, we demonstrate that nearly complete hybrid male sterility in Mimulus results from a simple genetic incompatibility between a single pair of heterospecific loci. We have genetically mapped this sterility effect: the M. guttatus allele at the hybrid male sterility 1 (hms1) locus acts dominantly in combination with recessive M. nasutus alleles at the hybrid male sterility 2 (hms2) locus to cause nearly complete hybrid male sterility. In a preliminary screen to find additional small-effect male sterility factors, we identified one additional locus that also contributes to some of the variation in hybrid male fertility. Interestingly, hms1 and hms2 also cause a significant reduction in hybrid female fertility, suggesting that sex-specific hybrid defects might share a common genetic basis. This possibility is supported by our discovery that recombination is reduced dramatically in a cross involving a parent with the hms1-hms2 incompatibility.     

Disruption of genetic interaction between two autosomal regions and the X chromosome causes reproductive isolation between mouse strains derived from different subspecies

Reproductive isolation that initiates speciation is likely caused by incompatibility among multiple loci in organisms belonging to genetically diverging populations. Laboratory C57BL/6J mice, which predominantly originated from Mus musculus domesticus, and a MSM/Ms strain derived from Japanese wild mice (M. m. molossinus, genetically close to M. m. musculus) are reproductively isolated. Their F1 hybrids are fertile, but successive intercrosses result in sterility. A consomic strain, C57BL/6J-ChrX(MSM), which carries the X chromosome of MSM/Ms in the C57BL/6J background, shows male sterility, suggesting a genetic incompatibility of the MSM/Ms X chromosome and other C57BL/6J chromosome(s). In this study, we conducted genomewide linkage analysis and subsequent QTL analysis using the sperm shape anomaly that is the major cause of the sterility of the C57BL/6J-ChrX(MSM) males. These analyses successfully detected significant QTL on chromosomes 1 and 11 that interact with the X chromosome. The introduction of MSM/Ms chromosomes 1 and 11 into the C57BL/6J-ChrX(MSM) background failed to restore the sperm-head shape, but did partially restore fertility. This result suggests that this genetic interaction may play a crucial role in the reproductive isolation between the two strains. A detailed analysis of the male sterility by intracytoplasmic sperm injection and zona-free in vitro fertilization demonstrated that the C57BL/6J-ChrX(MSM) spermatozoa have a defect in penetration through the zona pellucida of eggs.