X-Linked Signature of Reproductive Isolation in Humans is Mirrored in a Howler Monkey Hybrid Zone.

Reproductive isolation is a fundamental step in speciation. While sex chromosomes have been linked to reproductive isolation in many model systems, including hominids, genetic studies of the contribution of sex chromosome loci to speciation for natural populations are relatively sparse. Natural hybrid zones can help identify genomic regions contributing to reproductive isolation, like hybrid incompatibility loci, since these regions exhibit reduced introgression between parental species. Here, we use a primate hybrid zone (Alouatta palliata × Alouatta pigra) to test for reduced introgression of X-linked SNPs compared to autosomal SNPs. To identify X-linked sequence in A. palliata, we used a sex-biased mapping approach with whole-genome re-sequencing data. We then used genomic cline analysis with reduced-representation sequence data for parental A. palliata and A. pigra individuals and hybrids (n = 88) to identify regions with non-neutral introgression. We identified ~26 Mb of non-repetitive, putatively X-linked genomic sequence in A. palliata, most of which mapped collinearly to the marmoset and human X chromosomes. We found that X-linked SNPs had reduced introgression and an excess of ancestry from A. palliata as compared to autosomal SNPs. One outlier region with reduced introgression overlaps a previously described "desert" of archaic hominin ancestry on the human X chromosome. These results are consistent with a large role for the X chromosome in speciation across animal taxa and further, suggest shared features in the genomic basis of the evolution of reproductive isolation in primates.

Multiple forms of selection shape reproductive isolation in a primate hybrid zone.

Speciation occurs when populations diverge and become reproductively isolated from each other. Natural selection is commonly accepted to play a large role in this process, and it has been widely assumed that reproductive isolation often results as a by-product of divergence driven by adaptation in allopatry. When such populations come into secondary contact, reinforcement can act to strengthen reproductive isolation, but the frequency and importance of this process are still unknown. Here, we explored genomic signatures of selection in allopatry and sympatry for loci associated with reproductive isolation using a natural primate hybrid zone. By analysing reduced-representation sequencing data, we quantified admixture and population structure across a howler monkey hybrid zone and examined the relationship between locus-specific differentiation and introgression. We detected extensive admixture that was mostly limited to the narrow contact zone. Loci with reduced introgression into the heterospecific genomic background (the pattern expected for loci associated with reproductive isolation due to selection against hybrids) were significantly more differentiated between allopatric parental populations than loci with neutral and increased introgression, supporting the hypothesis that reproductive isolation is a by-product of divergence in allopatry. Further, loci with reduced introgression showed greater differentiation in sympatry than in allopatry, suggesting a role for reinforcement. Thus, our results reflect multiple forms of selection that have shaped reproductive isolation in this system. We conclude that reproductive isolation may have initially been driven by divergence in allopatry, but later reinforced by divergent selection in sympatry.

Effect of coffee agriculture management on the population structure of a forest dwelling rodent (Heteromys desmarestianus goldmani).

Most of the natural habitat in tropical regions exists as scattered fragments embedded in a matrix of different agricultural uses. As a result of this agricultural expansion, habitat loss and fragmentation have become the main drivers of biodiversity loss. Understanding the long-term effects of agricultural management on populations is of great importance for the development of successful conservation strategies. Our study uses genetic data to determine the effect of agricultural management practices on the population structure of a common tropical forest rodent (Heteromys desmarestianus goldmani). We sampled 136 individuals from one forest fragment and three coffee farms representing varying degrees of management intensity in southern Mexico. Using microsatellite markers, we evaluated the genetic structure of H. d. goldmani in the study area. Our results show higher genetic differentiation and lower connectivity for individuals within high and medium intensity coffee farms than for those near and within the forest fragments. Our results suggest that the population structure observed is driven by landscape characteristics other than distance.

Phylogenomic Insights into Mouse Evolution Using a Pseudoreference Approach.

Comparative genomic studies are now possible across a broad range of evolutionary timescales, but the generation and analysis of genomic data across many different species still present a number of challenges. The most sophisticated genotyping and down-stream analytical frameworks are still predominantly based on comparisons to high-quality reference genomes. However, established genomic resources are often limited within a given group of species, necessitating comparisons to divergent reference genomes that could restrict or bias comparisons across a phylogenetic sample. Here, we develop a scalable pseudoreference approach to iteratively incorporate sample-specific variation into a genome reference and reduce the effects of systematic mapping bias in downstream analyses. To characterize this framework, we used targeted capture to sequence whole exomes (∼54 Mbp) in 12 lineages (ten species) of mice spanning the Mus radiation. We generated whole exome pseudoreferences for all species and show that this iterative reference-based approach improved basic genomic analyses that depend on mapping accuracy while preserving the associated annotations of the mouse reference genome. We then use these pseudoreferences to resolve evolutionary relationships among these lineages while accounting for phylogenetic discordance across the genome, contributing an important resource for comparative studies in the mouse system. We also describe patterns of genomic introgression among lineages and compare our results to previous studies. Our general approach can be applied to whole or partitioned genomic data and is easily portable to any system with sufficient genomic resources, providing a useful framework for phylogenomic studies in mice and other taxa.

Functional organization of the genome may shape the species boundary in the house mouse.

Genomic features such as rate of recombination and differentiation have been suggested to play a role in species divergence. However, the relationship of these phenomena to functional organization of the genome in the context of reproductive isolation remains unexplored. Here, we examine genomic characteristics of the species boundaries between two house mouse subspecies (Mus musculus musculus/M. m. domesticus). These taxa form a narrow semipermeable zone of secondary contact across Central Europe. Due to the incomplete nature of reproductive isolation, gene flow in the zone varies across the genome. We present an analysis of genomic differentiation, rate of recombination, and functional composition of genes relative to varying amounts of introgression. We assessed introgression using 1,316 autosomal single nucleotide polymorphism markers, previously genotyped in hybrid populations from three transects. We found a significant relationship between amounts of introgression and both genomic differentiation and rate of recombination with genomic regions of reduced introgression associated with higher genomic differentiation and lower rates of recombination, and the opposite for genomic regions of extensive introgression. We also found a striking functional polarization of genes based on where they are expressed in the cell. Regions of elevated introgression exhibit a disproportionate number of genes involved in signal transduction functioning at the cell periphery, among which olfactory receptor genes were found to be the most prominent group. Conversely, genes expressed intracellularly and involved in DNA binding were the most prevalent in regions of reduced introgression. We hypothesize that functional organization of the genome is an important driver of species divergence.

The uncharacterized gene 1700093K21Rik and flanking regions are correlated with reproductive isolation in the house mouse, Mus musculus.

Reproductive barriers exist between the house mouse subspecies, Mus musculus musculus and M. m. domesticus, members of the Mus musculus species complex, primarily as a result of hybrid male infertility, and a hybrid zone exists where their ranges intersect in Europe. Using single nucleotide polymorphisms (SNPs) diagnostic for the two taxa, the extent of introgression across the genome was previously compared in these hybrid populations. Sixty-nine of 1316 autosomal SNPs exhibited reduced introgression in two hybrid zone transects suggesting maladaptive interactions among certain loci. One of these markers is within a region on chromosome 11 that, in other studies, has been associated with hybrid male sterility of these subspecies. We assessed sequence variation in a 20 Mb region on chromosome 11 flanking this marker, and observed its inclusion within a roughly 150 kb stretch of DNA showing elevated sequence differentiation between the two subspecies. Four genes are associated with this genomic subregion, with two entirely encompassed. One of the two genes, the uncharacterized 1700093K21Rik gene, displays distinguishing features consistent with a potential role in reproductive isolation between these subspecies. Along with its expression specifically within spermatogenic cells, we present various sequence analyses that demonstrate a high rate of molecular evolution of this gene, as well as identify a subspecies amino acid variant resulting in a structural difference. Taken together, the data suggest a role for this gene in reproductive isolation.

Sperm-related phenotypes implicated in both maintenance and breakdown of a natural species barrier in the house mouse.

The house mouse hybrid zone (HMHZ) is a species barrier thought to be maintained by a balance between dispersal and natural selection against hybrids. While the HMHZ is characterized by frequency discontinuities for some sex chromosome markers, there is an unexpected large-scale regional introgression of a Y chromosome across the barrier, in defiance of Haldane's rule. Recent work suggests that a major force maintaining the species barrier acts through sperm traits. Here, we test whether the Y chromosome penetration of the species barrier acts through sperm traits by assessing sperm characteristics of wild-caught males directly in a field laboratory set up in a Y introgression region of the HMHZ, later calculating the hybrid index of each male using 1401 diagnostic single nucleotide polymorphisms (SNPs). We found that both sperm count (SC) and sperm velocity were significantly reduced across the natural spectrum of hybrids. However, SC was more than rescued in the presence of the invading Y. Our results imply an asymmetric advantage for Y chromosome introgression consistent with the observed large-scale introgression. We suggest that selection on sperm-related traits probably explains a large component of patterns observed in the natural hybrid zone, including the Y chromosome penetration.

Genome-wide architecture of reproductive isolation in a naturally occurring hybrid zone between Mus musculus musculus and M. m. domesticus.

Studies of a hybrid zone between two house mouse subspecies (Mus musculus musculus and M. m. domesticus) along with studies using laboratory crosses reveal a large role for the X chromosome and multiple autosomal regions in reproductive isolation as a consequence of disrupted epistasis in hybrids. One limitation of previous work has been that most of the identified genomic regions have been large. The goal here is to detect and characterize precise genomic regions underlying reproductive isolation. We surveyed 1401 markers evenly spaced across the genome in 679 mice collected from two different transects. Comparisons between transects provide a means for identifying common patterns that likely reflect intrinsic incompatibilities. We used a genomic cline approach to identify patterns that correspond to epistasis. From both transects, we identified contiguous regions on the X chromosome in which markers were inferred to be involved in epistatic interactions. We then searched for autosomal regions showing the same patterns and found they constitute about 5% of autosomal markers. We discovered substantial overlap between these candidate regions underlying reproductive isolation and QTL for hybrid sterility identified in laboratory crosses. Analysis of gene content in these regions suggests a key role for several mechanisms, including the regulation of transcription, sexual conflict and sexual selection operating at both the postmating prezygotic and postzygotic stages of reproductive isolation. Taken together, these results indicate that speciation in two recently diverged (c. 0.5 Ma) house mouse subspecies is complex, involving many genes dispersed throughout the genome and associated with distinct functions.

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 F1 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 F1 autosomal background, and we measured a suite of male reproductive traits. Reproductive deficits were observed in most F1 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 F1 male sterility appears to be caused mainly by X-autosome incompatibilities, X-Y incompatibilities contribute to some aspects of sterility.

Adaptive evolution and effective population size in wild house mice.

Estimates of the proportion of amino acid substitutions that have been fixed by selection (α) vary widely among taxa, ranging from zero in humans to over 50% in Drosophila. This wide range may reflect differences in the efficacy of selection due to differences in the effective population size (N(e)). However, most comparisons have been made among distantly related organisms that differ not only in N(e) but also in many other aspects of their biology. Here, we estimate α in three closely related lineages of house mice that have a similar ecology but differ widely in N(e): Mus musculus musculus (N(e) ∼ 25,000-120,000), M. m. domesticus (N(e) ∼ 58,000-200,000), and M. m. castaneus (N(e) ∼ 200,000-733,000). Mice were genotyped using a high-density single nucleotide polymorphism array, and the proportions of replacement and silent mutations within subspecies were compared with those fixed between each subspecies and an outgroup, Mus spretus. There was significant evidence of positive selection in M. m. castaneus, the lineage with the largest N(e), with α estimated to be approximately 40%. In contrast, estimates of α for M. m. domesticus (α = 13%) and for M. m. musculus (α = 12 %) were much smaller. Interestingly, the higher estimate of α for M. m. castaneus appears to reflect not only more adaptive fixations but also more effective purifying selection. These results support the hypothesis that differences in N(e) contribute to differences among species in the efficacy of selection.

Measures of linkage disequilibrium among neighbouring SNPs indicate asymmetries across the house mouse hybrid zone.

Theory predicts that naturally occurring hybrid zones between genetically distinct taxa can move over space and time as a result of selection and/or demographic processes, with certain types of hybrid zones being more or less likely to move. Determining whether a hybrid zone is stationary or moving has important implications for understanding evolutionary processes affecting interactions in hybrid populations. However, direct observations of hybrid zone movement are difficult to make unless the zone is moving rapidly. Here, evidence for movement in the house mouse Mus musculus domesticus × Mus musculus musculus hybrid zone is provided using measures of LD and haplotype structure among neighbouring SNP markers from across the genome. Local populations of mice across two transects in Germany and the Czech Republic were sampled, and a total of 1301 mice were genotyped at 1401 markers from the nuclear genome. Empirical measures of LD provide evidence for extinction and (re)colonization in single populations and, together with simulations, suggest hybrid zone movement because of either geography-dependent asymmetrical dispersal or selection favouring one subspecies over the other.

Reinforcement selection acting on the European house mouse hybrid zone.

Behavioural isolation may lead to complete speciation when partial postzygotic isolation acts in the presence of divergent-specific mate-recognition systems. These conditions exist where Mus musculus musculus and M. m. domesticus come into contact and hybridize. We studied two mate-recognition signal systems, based on urinary and salivary proteins, across a Central European portion of the mouse hybrid zone. Introgression of the genomic regions responsible for these signals: the major urinary proteins (MUPs) and androgen binding proteins (ABPs), respectively, was compared to introgression at loci assumed to be nearly neutral and those under selection against hybridization. The preference of individuals taken from across the zone regarding these signals was measured in Y mazes, and we develop a model for the analysis of the transition of such traits under reinforcement selection. The strongest assortative preferences were found in males for urine and females for ABP. Clinal analyses confirm nearly neutral introgression of an Abp locus and two loci closely linked to the Abp gene cluster, whereas two markers flanking the Mup gene region reveal unexpected introgression. Geographic change in the preference traits matches our reinforcement selection model significantly better than standard cline models. Our study confirms that behavioural barriers are important components of reproductive isolation between the house mouse subspecies.

The variable genomic architecture of isolation between hybridizing species of house mice.

Studies of the genetics of hybrid zones can provide insight into the genomic architecture of species boundaries. By examining patterns of introgression of multiple loci across a hybrid zone, it may be possible to identify regions of the genome that have experienced selection. Here, we present a comparison of introgression in two replicate transects through the house mouse hybrid zone through central Europe, using data from 41 single nucleotide markers. Using both genomic and geographic clines, we found many differences in patterns of introgression between the two transects, as well as some similarities. We found that many loci may have experienced the effects of selection at linked sites, including selection against hybrid genotypes, as well as positive selection in the form of genotypes introgressed into a foreign genetic background. We also found many positive associations of conspecific alleles among unlinked markers, which could be caused by epistatic interactions. Different patterns of introgression in the two transects highlight the challenge of using hybrid zones to identify genes underlying isolation and raise the possibility that the genetic basis of isolation between these species may be dependent on the local population genetic make-up or the local ecological setting.

The endothelial-specific regulatory mutation, Mvwf1, is a common mouse founder allele.

Mvwf1 is a cis-regulatory mutation previously identified in the RIIIS/J mouse strain that causes a unique tissue-specific switch in the expression of an N-acetylgalactosaminyltransferase, B4GALNT2, from intestinal epithelium to vascular endothelium. Vascular B4galnt2 expression results in aberrant glycosylation of von Willebrand Factor (VWF) and accelerated VWF clearance from plasma. We now report that 13 inbred mouse strains share the Mvwf1 tissue-specific switch and low VWF phenotype, including five wild-derived strains. Genomic sequencing identified a highly conserved 97-kb Mvwf1 haplotype block shared by these strains that encompasses a 30-kb region of high nucleotide sequence divergence from C57BL6/J flanking B4galnt2 exon 1. The analysis of a series of bacterial artificial chromosome (BAC) transgenes containing B4galnt2 derived from the RIIIS/J or C57BL6/J inbred mouse strains demonstrates that the corresponding sequences are sufficient to confer the vessel (RIIIS/J) or intestine (C57BL6/J)-specific expression patterns. Taken together, our data suggest that the region responsible for the Mvwf1 regulatory switch lies within an approximately 30-kb genomic interval upstream of the B4galnt2 gene. The observation that Mvwf1 is present in multiple wild-derived strains suggests that this locus may be retained in wild mouse populations due to positive selection. Similar selective pressures could contribute to the high prevalence of von Willebrand disease in humans.

Genome-wide patterns of gene flow across a house mouse hybrid zone.

Hybrid zones between closely related species or subspecies provide useful settings for studying the genetic architecture of speciation. Using markers distributed throughout the mouse genome, we use a hybrid zone between two recently diverged species of house mice (Mus musculus and Mus domesticus) as a natural mapping experiment to identify genomic regions that may be involved in reproductive isolation. Using cline analysis we document a nearly 50-fold variation in level of introgression among markers. Some markers have extremely narrow cline widths; these genomic regions may contribute to reproductive isolation. Biological processes associated with these narrow clines include physiological and immune responses to the environment as well as physiological and behavioral aspects of reproduction. Other autosomal markers exhibit asymmetrically broad clines, usually with high frequencies of M. domesticus alleles on the M. musculus side of the hybrid zone. These markers identify genome regions likely housing genes with alleles that are spreading from one species to the other. Biological processes associated with these wide clines include cell signaling, olfaction, and pheromone response. These processes play important roles in survival and reproduction, and associated genes are likely targets of selection. Patterns of linkage disequilibrium in the center of the hybrid zone suggest that isolation may be caused by multiple epistatic interactions between sets of genes. These data highlight the complex genetic architecture underlying speciation even at early stages of divergence and point to some of the biological processes that may govern this architecture.

Inefficient purifying selection: the mammalian Y chromosome in the rodent genus Mus.

Two related genes with potentially similar functions, one on the Y chromosome and one on the X chromosome, were examined to determine if they evolved differently because of their chromosomal positions. Six hundred fifty-seven base pairs of coding sequence of Jarid1d (Smcy) on the Y chromosome and Jarid1c (Smcx) on the X chromosome were sequenced in 13 rodent taxa. An analysis of replacement and silent substitutions, using a counting method designed for samples with small evolutionary distances, showed a significant difference between the two genes. The different patterns of replacement and silent substitutions within Jarid1d and Jarid1c may be a result of evolutionary mechanisms that are particularly strong on the Y chromosome because of its unique properties. These findings are similar to results of previous studies of Y chromosomal genes in these and other mammalian taxa, suggesting that genes on the mammalian Y evolve in a chromosome-specific manner.

Male-driven evolution in closely related species of the mouse genus Mus.

Recently, other researchers have found that closely related primate species had a lower male-to-female mutation rate ratio (alpha) than distantly related species. To determine if this is a general phenomenon affecting other mammalian orders, eleven species or subspecies of the rodent genus Mus and two outgroup species were compared. Intron sequences from a gene in the nonrecombining region of the Y chromosome Jarid1d (Smcy) and its X chromosomal gametolog, Jarid1c (Smcx), were analyzed in a phylogenetic context. The male-to-female mutation rate ratio for all thirteen taxa is approximately 2.5, which is similar to previous estimates in more distantly related rodents. However, when branches with lengths of more than 2.5% were removed from the analysis, the male-to-female mutation rate ratio dropped to 0.9. Thus, in closely related rodents, as in closely related primates, the male-to-female mutation rate ratio is lower than expected.

Positive selection for indel substitutions in the rodent sperm protein catsper1.

Catsper1 is a voltage-gated calcium channel located in the plasma membrane of the sperm tail and is necessary for sperm motility and fertility in mice. We here examine the evolutionary pattern of Catsper1 from nine species of the rodent subfamily Murinae of family Muridae. We show that the rate of insertion/deletion (indel) substitutions in exon 1 of the gene is 4-15 times that in introns or neutral genomic regions, suggesting the presence of strong positive selection that promotes fixations of indel mutations in exon 1. The number of indel polymorphisms within species appears higher than expected from interspecific comparisons, although there are too little data to provide a statistically significant conclusion. These results, together with an earlier report in primates, indicate that positive selection promoting length variation in Catsper1 may be widespread in mammals. A structural model of Catsper1 suggested the importance of the exon 1-encoded region in regulating channel inactivation, which may affect sperm mobility and sperm competition. Our findings provide a necessary foundation for future experimental investigations of Catsper1's function in sperm physiology and role in sperm competition using rodent models.

Evolutionary strata on the mouse X chromosome correspond to strata on the human X chromosome.

Lahn and Page previously observed that genes on the human X chromosome were physically arranged along the chromosome in "strata," roughly ordered by degree of divergence from related genes on the Y chromosome. They hypothesized that this ordering results from a historical series of suppressions of recombination along the mammalian Y chromosome, thereby allowing formerly recombining X and Y chromosomal genes to diverge independently. Here predictions of this hypothesis are confirmed in a nonprimate mammalian order, Rodentia, through an analysis of eight gene pairs from the X and Y chromosomes of the house mouse, Mus musculus. The mouse X chromosome has been rearranged relative to the human X, so strata were not found in the same physical order on the mouse X. However, based on synonymous evolutionary distances, X-linked genes in M. musculus fall into the same strata as orthologous genes in humans, as predicted. The boundary between strata 2 and 3 is statistically significant, but the boundary between strata 1 and 2 is not significant in mice. An analysis of smaller fragments of Smcy, Smcx, Zfy, and Zfx from seven species of Mus confirmed that the strata in Mus musculus were representative of the genus Mus.

Differential rates of evolution for the ZFY-related zinc finger genes, Zfy, Zfx, and Zfa in the mouse genus Mus.

A comparative study of the last exon of the zinc finger genes Zfx, Zfy, and Zfa from species of mice in the genus Mus was conducted to assess the extent of gene-specific and chromosome-specific effects on the evolutionary patterns among related X-, Y-, and autosomal-linked genes. Phylogenetic analyses of 29 sequences from Zfx, Zfa, and Zfy from 10 taxa were performed to infer relatedness among the zinc finger loci, and codon-based maximum likelihood analyses were conducted to assess evolutionary pattern among genes. Five models of nucleotide sequence evolution were applied and compared using a likelihood ratio test. Estimates of nonsynonymous to synonymous changes (dN/dS) for these genes suggest that amino acid substitutions are occurring at a more rapid rate across the autosomal- and Y-specific lineages compared to the X-specific lineage, with the Y-specific lineage showing the highest rate under certain models. The data suggest the action of gene-specific effects on evolutionary pattern. In particular, Zfa and Zfy genes, both with presumed restricted expression, appear less functionally constrained relative to ubiquitously expressed Zfx. Slightly elevated dN/dS for Zfy genes in comparison to Zfa also suggest Y-specific effects.