Heterogeneous Histories of Recombination Suppression on Stickleback Sex Chromosomes

How consistent are the evolutionary trajectories of sex chromosomes shortly after they form? Insights into the evolution of recombination, differentiation, and degeneration can be provided by comparing closely related species with homologous sex chromosomes. The sex chromosomes of the threespine stickleback (Gasterosteus aculeatus) and its sister species, the Japan Sea stickleback (G. nipponicus), have been well characterized. Little is known, however, about the sex chromosomes of their congener, the blackspotted stickleback (G. wheatlandi). We used pedigrees to obtain experimentally phased whole genome sequences from blackspotted stickleback X and Y chromosomes. Using multispecies gene trees and analysis of shared duplications, we demonstrate that Chromosome 19 is the ancestral sex chromosome and that its oldest stratum evolved in the common ancestor of the genus. After the blackspotted lineage diverged, its sex chromosomes experienced independent and more extensive recombination suppression, greater X–Y differentiation, and a much higher rate of Y degeneration than the other two species. These patterns may result from a smaller effective population size in the blackspotted stickleback. A recent fusion between the ancestral blackspotted stickleback Y chromosome and Chromosome 12, which produced a neo-X and neo-Y, may have been favored by the very small size of the recombining region on the ancestral sex chromosome. We identify six strata on the ancestral and neo-sex chromosomes where recombination between the X and Y ceased at different times. These results confirm that sex chromosomes can evolve large differences within and between species over short evolutionary timescales.     

The Epigenome of Evolving Drosophila Neo-Sex Chromosomes: Dosage Compensation and Heterochromatin Formation

Sex chromosomes originated from autosomes but have evolved a highly specialized chromatin structure. Drosophila Y chromosomes are composed entirely of silent heterochromatin, while male X chromosomes have highly accessible chromatin and are hypertranscribed as a result of dosage compensation. Here, we dissect the molecular mechanisms and functional pressures driving heterochromatin formation and dosage compensation of the recently formed neo-sex chromosomes of Drosophila miranda. We show that the onset of heterochromatin formation on the neo-Y is triggered by an accumulation of repetitive DNA. The neo-X has evolved partial dosage compensation and we find that diverse mutational paths have been utilized to establish several dozen novel binding consensus motifs for the dosage compensation complex on the neo-X, including simple point mutations at pre-binding sites, insertion and deletion mutations, microsatellite expansions, or tandem amplification of weak binding sites. Spreading of these silencing or activating chromatin modifications to adjacent regions results in massive mis-expression of neo-sex linked genes, and little correspondence between functionality of genes and their silencing on the neo-Y or dosage compensation on the neo-X. Intriguingly, the genomic regions being targeted by the dosage compensation complex on the neo-X and those becoming heterochromatic on the neo-Y show little overlap, possibly reflecting different propensities along the ancestral chromosome that formed the sex chromosome to adopt active or repressive chromatin configurations. Our findings have broad implications for current models of sex chromosome evolution, and demonstrate how mechanistic constraints can limit evolutionary adaptations. Our study also highlights how evolution can follow predictable genetic trajectories, by repeatedly acquiring the same 21-bp consensus motif for recruitment of the dosage compensation complex, yet utilizing a diverse array of random mutational changes to attain the same phenotypic outcome.     

Sequence differentiation associated with an inversion on the neo-X chromosome of Drosophila americana

Sex chromosomes originate from pairs of autosomes that acquire controlling genes in the sex-determining cascade. Universal mechanisms apparently influence the evolution of sex chromosomes, because this chromosomal pair is characteristically heteromorphic in a broad range of organisms. To examine the pattern of initial differentiation between sex chromosomes, sequence analyses were performed on a pair of newly formed sex chromosomes in Drosophila americana. This species has neo-sex chromosomes as a result of a centromeric fusion between the X chromosome and an autosome. Sequences were analyzed from the Alcohol dehydrogenase (Adh), big brain (bib), and timeless (tim) gene regions, which represent separate positions along this pair of neo-sex chromosomes. In the northwestern range of the species, the bib and Adh regions exhibit significant sequence differentiation for neo-X chromosomes relative to neo-Y chromosomes from the same geographic region and other chromosomal populations of D. americana. Furthermore, a nucleotide site defining a common haplotype in bib is shown to be associated with a paracentric inversion [In(4)ab] on the neo-X chromosome, and this inversion suppresses recombination between neo-X and neo-Y chromosomes. These observations are consistent with the inversion acting as a recombination modifier that suppresses exchange between these neo-sex chromosomes, as predicted by models of sex chromosome evolution.     

Sex Chromosome Turnover Contributes to Genomic Divergence between Incipient Stickleback Species

Sex chromosomes turn over rapidly in some taxonomic groups, where closely related species have different sex chromosomes. Although there are many examples of sex chromosome turnover, we know little about the functional roles of sex chromosome turnover in phenotypic diversification and genomic evolution. The sympatric pair of Japanese threespine stickleback (Gasterosteus aculeatus) provides an excellent system to address these questions: the Japan Sea species has a neo-sex chromosome system resulting from a fusion between an ancestral Y chromosome and an autosome, while the sympatric Pacific Ocean species has a simple XY sex chromosome system. Furthermore, previous quantitative trait locus (QTL) mapping demonstrated that the Japan Sea neo-X chromosome contributes to phenotypic divergence and reproductive isolation between these sympatric species. To investigate the genomic basis for the accumulation of genes important for speciation on the neo-X chromosome, we conducted whole genome sequencing of males and females of both the Japan Sea and the Pacific Ocean species. No substantial degeneration has yet occurred on the neo-Y chromosome, but the nucleotide sequence of the neo-X and the neo-Y has started to diverge, particularly at regions near the fusion. The neo-sex chromosomes also harbor an excess of genes with sex-biased expression. Furthermore, genes on the neo-X chromosome showed higher non-synonymous substitution rates than autosomal genes in the Japan Sea lineage. Genomic regions of higher sequence divergence between species, genes with divergent expression between species, and QTL for inter-species phenotypic differences were found not only at the regions near the fusion site, but also at other regions along the neo-X chromosome. Neo-sex chromosomes can therefore accumulate substitutions causing species differences even in the absence of substantial neo-Y degeneration.     

Gene content evolution on the X chromosome

Compared with autosomes, the X chromosome shows different patterns of evolution as a result of its hemizygosity in males. Additionally, inactivation of the X during spermatogenesis can make the X chromosome an unfavorable location for male-specific genes. These factors can help to explain why in many species gene content of the X chromosome differs from that of autosomes. Indeed, the X chromosome in mouse is enriched for male-specific genes while they are depleted on the X in Drosophila but show neither of these trends in mosquito. Here, we will discuss recent findings on the ancestral and neo-X chromosomes in Drosophila that support sexual antagonism as a force shaping gene content evolution of sex chromosomes and suggest that selection could be driving male-biased genes off the X.     

A dynamic view of sex chromosome evolution

Sex chromosomes are derived from ordinary autosomes. The X chromosome is thought to maintain most of its ancestral genes over evolutionary time, whereas its Y counterpart degenerates, owing to its lack of recombination. Genomic analyses of young sex chromosome pairs support this view and have shed light on the evolutionary processes underlying loss of gene function on the Y. Studies of ancestral sex chromosomes, however, have also revealed that the process of sex chromosome evolution can be more dynamic than traditionally appreciated. In particular, ancient Y-chromosomes are characterized not only by a loss of genes relative to the X but also by recurrent gains of individual genes or genomic regions, and they often accumulate genes beneficial to males. Furthermore, X chromosomes are not passive players in this evolutionary process but respond both to their sex-biased transmission and to Y-chromosome degeneration, through feminization and the evolution of dosage compensation.     

A selective sweep associated with a recent gene transposition in Drosophila miranda

In Drosophila miranda, a chromosome fusion between the Y chromosome and the autosome corresponding to Muller's element C has created a new sex chromosome system. The chromosome attached to the ancestral Y chromosome is transmitted paternally and hence is not exposed to crossing over. This chromosome, conventionally called the neo-Y, and the homologous neo-X chromosome display many properties of evolving sex chromosomes. We report here the transposition of the exuperantia1 (exu1) locus from a neo-sex chromosome to the ancestral X chromosome of D. miranda. Exu1 is known to have several critical developmental functions, including a male-specific role in spermatogenesis. The ancestral location of exu1 is conserved in the sibling species of D. miranda, as well as in a more distantly related species. The transposition of exu1 can be interpreted as an adaptive fixation, driven by a selective advantage conferred by its effect on dosage compensation. This explanation is supported by the pattern of within-species sequence variation at exu1 and the nearby exu2 locus. The implications of this phenomenon for genome evolution are discussed.     

Contrasting patterns of molecular evolution of the genes on the new and old sex chromosomes of Drosophila miranda

In organisms with chromosomal sex determination, sex is determined by a set of dimorphic sex chromosomes that are thought to have evolved from a set of originally homologous chromosomes. The chromosome inherited only through the heterogametic sex (the Y chromosome in the case of male heterogamety) often exhibits loss of genetic activity for most of the genes carried on its homolog and is hence referred to as degenerate. The process by which the proto-Y chromosome loses its genetic activity has long been the subject of much speculation. We present a DNA sequence variation analysis of marker genes on the evolving sex chromosomes (neo-sex chromosomes) of Drosophila miranda. Due to its relatively recent origin, the neo-Y chromosome of this species is presumed to be still experiencing the forces responsible for the loss of its genetic activity. Indeed, several previous studies have confirmed the presence of some active loci on this chromosome. The genes on the neo-Y chromosome surveyed in the current study show generally lower levels of variation compared with their counterparts on the neo-X chromosome or an X-linked gene. This is in accord with a reduced effective population size of the neo-Y chromosome. Interestingly, the rate of replacement nucleotide substitutions for the neo-Y linked genes is significantly higher than that for the neo-X linked genes. This is not expected under a model where the faster evolution of the X chromosome is postulated to be the main force driving the degeneration of the Y chromosome.     

Positive selection near an inversion breakpoint on the neo-X chromosome of Drosophila americana

Unique features of heteromorphic sex chromosomes are produced as a consequence of sex-linked transmission. Alternative models concerning the evolution of sex chromosomes can be classified in terms of genetic drift or positive selection being the primary mechanism of divergence between this chromosomal pair. This study examines early changes on a newly acquired chromosomal arm of the X in Drosophila americana, which was derived from a centromeric fusion between the ancestral X and previously autosomal chromosome 4 (element B). Breakpoints of a chromosomal inversion In(4)a, which is restricted to the neo-X, are identified and used to guide a sequence analysis along chromosome 4. Loci flanking the distal breakpoint exhibit patterns of sequence diversity consistent with neutral evolution, yet loci near the proximal breakpoint reveal distinct imprints of positive selection within the neo-X chromosomal class containing In(4)a. Data from six separate positions examined throughout the proximal region reveal a pattern of recent turnover driven by two independent sweeps among chromosomes with the inverted gene arrangement. Selection-mediated establishment of an extended haplotype associated with recombination-suppressing inversions on the neo-X indicates a pattern of active coadaptation apparently initiated by X-linked transmission and potentially sustained by intralocus sexual conflict.     

Patterns of replication in the neo-sex chromosomes ofDrosophila nasuta albomicans

Drosophila nasuta albomicans (with 2n = 6), contains a pair of metacentric neo-sex chromosomes. Phylogenetically these are products of centric fusion between ancestral sex (X, Y) chromosomes and an autosome (chromosome 3). The polytene chromosome complement of males with a neo-X- and neo-Y-chromosomes has revealed asynchrony in replication between the two arms of the neo-sex chromosomes. The arm which represents the ancestral X-chromosome is faster replicating than the arm which represents ancestral autosome. The latter arm of the neo-sex chromosome is synchronous with other autosomes of the complement. We conclude that one arm of the neo-X/Y is still mimicking the features of an autosome while the other arm has the features of a classical X/Y-chromosome. This X-autosome translocation differs from the other evolutionary X-autosome translocations known in certain species ofDrosophila.     

Genomic degradation of a young Y chromosome in Drosophila miranda

Background

Y chromosomes are derived from ordinary autosomes and degenerate because of a lack of recombination. Well-studied Y chromosomes only have few of their original genes left and contain little information about their evolutionary origin. Here, we take advantage of the recently formed neo-Y chromosome of Drosophila miranda to study the processes involved in Y degeneration on a genomic scale.

Results

We obtained sequence information from 14 homologous bacterial artificial chromosome (BAC) clones from the neo-X and neo-Y chromosome of D. miranda, encompassing over 2.5 Mb of neo-sex-linked DNA. A large fraction of neo-Y DNA is composed of repetitive and transposable-element-derived DNA (20% of total DNA) relative to their homologous neo-X linked regions (1%). The overlapping regions of the neo-sex linked BAC clones contain 118 gene pairs, half of which are pseudogenized on the neo-Y. Pseudogenes evolve significantly faster on the neo-Y than functional genes, and both functional and non-functional genes show higher rates of protein evolution on the neo-Y relative to their neo-X homologs. No heterogeneity in levels of degeneration was detected among the regions investigated. Functional genes on the neo-Y are under stronger evolutionary constraint on the neo-X, but genes were found to degenerate randomly on the neo-Y with regards to their function or sex-biased expression patterns.

Conclusion

Patterns of genome evolution in D. miranda demonstrate that degeneration of a recently formed Y chromosome can proceed very rapidly, by both an accumulation of repetitive DNA and degeneration of protein-coding genes. Our data support a random model of Y inactivation, with little heterogeneity in degeneration among genomic regions, or between functional classes of genes or genes with sex-biased expression patterns.     

Reduced levels of microsatellite variability on the neo-Y chromosome of Drosophila miranda

BACKGROUND: In many species, sex is determined by a system involving X and Y chromosomes, the latter having lost much of their genetic activity. Sex chromosomes have evolved independently many times, and several different mechanisms responsible for the degeneration of the Y chromosome have been proposed. Here, we have taken advantage of the secondary sex chromosome pair in Drosophila miranda to test for the effects of evolutionary forces involved in the early stages of Y-chromosome degeneration. Because of a fusion of one of the autosomes to the Y chromosome, a neo-Y chromosome and a neo-X chromosome have been formed, resulting in the transmission of formerly autosomal genes in association with the sex chromosomes. RESULTS: We found a 25-fold lower level of variation at microsatellites located on the neo-Y chromosome compared with homologous loci on the neo-X chromosome, or with autosomal and X-linked microsatellites. Sequence analyses of the region flanking the microsatellites suggested that the neo-sex chromosomes originated about 1 million years ago. CONCLUSIONS: Variability of the neo-Y chromosome of D. miranda is substantially reduced below expectations at mutation-drift equilibrium. Such a reduction is predicted by theories of the degeneration of the Y chromosome. Another possibility is that there is little or no mutation at microsatellite loci on a non-recombining chromosome such as the neo-Y, but this seems inconsistent with other data.     

Natural variation of the Y chromosome suppresses sex ratio distortion and modulates testis-specific gene expression in Drosophila simulans

X-linked sex-ratio distorters that disrupt spermatogenesis can cause a deficiency in functional Y-bearing sperm and a female-biased sex ratio. Y-linked modifiers that restore a normal sex ratio might be abundant and favored when a X-linked distorter is present. Here we investigated natural variation of Y-linked suppressors of sex-ratio in the Winters systems and the ability of these chromosomes to modulate gene expression in Drosophila simulans. Seventy-eight Y chromosomes of worldwide origin were assayed for their resistance to the X-linked sex-ratio distorter gene Dox. Y chromosome diversity caused males to sire ∼63% to ∼98% female progeny. Genome-wide gene expression analysis revealed hundreds of genes differentially expressed between isogenic males with sensitive (high sex ratio) and resistant (low sex ratio) Y chromosomes from the same population. Although the expression of about 75% of all testis-specific genes remained unchanged across Y chromosomes, a subset of post-meiotic genes was upregulated by resistant Y chromosomes. Conversely, a set of accessory gland-specific genes and mitochondrial genes were downregulated in males with resistant Y chromosomes. The D. simulans Y chromosome also modulated gene expression in XXY females in which the Y-linked protein-coding genes are not transcribed. The data suggest that the Y chromosome might exert its regulatory functions through epigenetic mechanisms that do not require the expression of protein-coding genes. The gene network that modulates sex ratio distortion by the Y chromosome is poorly understood, other than that it might include interactions with mitochondria and enriched for genes expressed in post-meiotic stages of spermatogenesis.     

Reduced sequence variability on the Neo-Y chromosome of Drosophila americana americana.

Sex chromosomes are generally morphologically and functionally distinct, but the evolutionary forces that cause this differentiation are poorly understood. Drosophila americana americana was used in this study to examine one aspect of sex chromosome evolution, the degeneration of nonrecombining Y chromosomes. The primary X chromosome of D. a. americana is fused with a chromosomal element that was ancestrally an autosome, causing this homologous chromosomal pair to segregate with the sex chromosomes. Sequence variation at the Alcohol Dehydrogenase (Adh) gene was used to determine the pattern of nucleotide variation on the neo-sex chromosomes in natural populations. Sequences of Adh were obtained for neo-X and neo-Y chromosomes of D. a. americana, and for Adh of D. a. texana, in which it is autosomal. No significant sequence differentiation is present between the neo-X and neo-Y chromosomes of D. a. americana or the autosomes of D. a. texana. There is a significantly lower level of sequence diversity on the neo-Y chromosome relative to the neo-X in D. a. americana. This reduction in variability on the neo-Y does not appear to have resulted from a selective sweep. Coalescent simulations of the evolutionary transition of an autosome into a Y chromosome indicate there may be a low level of recombination between the neo-X and neo-Y alleles of Adh and that the effective population size of this chromosome may have been reduced below the expected value of 25% of the autosomal effective size, possibly because of the effects of background selection or sexual selection.     

Evolution and Survival on Eutherian Sex Chromosomes

Since the two eutherian sex chromosomes diverged from an ancestral autosomal pair, the X has remained relatively gene-rich, while the Y has lost most of its genes through the accumulation of deleterious mutations in nonrecombining regions. Presently, it is unclear what is distinctive about genes that remain on the Y chromosome, when the sex chromosomes acquired their unique evolutionary rates, and whether X-Y gene divergence paralleled that of paralogs located on autosomes. To tackle these questions, here we juxtaposed the evolution of X and Y homologous genes (gametologs) in eutherian mammals with their autosomal orthologs in marsupial and monotreme mammals. We discovered that genes on the X and Y acquired distinct evolutionary rates immediately following the suppression of recombination between the two sex chromosomes. The Y-linked genes evolved at higher rates, while the X-linked genes maintained the lower evolutionary rates of the ancestral autosomal genes. These distinct rates have been maintained throughout the evolution of X and Y. Specifically, in humans, most X gametologs and, curiously, also most Y gametologs evolved under stronger purifying selection than similarly aged autosomal paralogs. Finally, after evaluating the current experimental data from the literature, we concluded that unique mRNA/protein expression patterns and functions acquired by Y (versus X) gametologs likely contributed to their retention. Our results also suggest that either the boundary between sex chromosome strata 3 and 4 should be shifted or that stratum 3 should be divided into two strata.     

The chicken (Gallus gallus) Z chromosome contains at least three nonlinear evolutionary strata

Birds have female heterogamety with Z and W sex chromosomes. These evolved from different autosomal precursor chromosomes than the mammalian X and Y. However, previous work has suggested that the pattern and process of sex chromosome evolution show many similarities across distantly related organisms. Here we show that stepwise restriction of recombination between the protosex chromosomes of birds has resulted in regions of the chicken Z chromosome showing discrete levels of divergence from W homologs (gametologs). The 12 genes analyzed fall into three levels of estimated divergence values, with the most recent divergence (d_S = 0.18–0.21) displayed by 6 genes in a region on the Z chromosome corresponding to the interval 1–11 Mb of the assembled genome sequence. Another 4 genes show intermediate divergence (d_S = 0.27–0.38) and are located in the interval 16–53 Mb. Two genes (at positions 42 and 50 Mb) with higher d_S values are located proximal to the most distal of the 4 genes with intermediate divergence, suggesting an inversion event. The distribution of genes and their divergence indicate at least three evolutionary strata, with estimated times for cessation of recombination between Z and W of 132–150 (stratum 1), 71–99 (stratum 2), and 47–57 (stratum 3) million years ago. An inversion event, or some other form of intrachromosomal rearrangement, subsequent to the formation of strata 1 and 2 has scrambled the gene order to give rise to the nonlinear arrangement of evolutionary strata currently seen on the chicken Z chromosome. These observations suggest that the progressive restriction of recombination is an integral feature of sex chromosome evolution and occurs also in systems of female heterogamety.     

Protein evolution and codon usage bias on the neo-sex chromosomes of Drosophila miranda

The neo-sex chromosomes of Drosophila miranda constitute an ideal system to study the effects of recombination on patterns of genome evolution. Due to a fusion of an autosome with the Y chromosome, one homolog is transmitted clonally. Here, I compare patterns of molecular evolution of 18 protein-coding genes located on the recombining neo-X and their homologs on the nonrecombining neo-Y chromosome. The rate of protein evolution has significantly increased on the neo-Y lineage since its formation. Amino acid substitutions are accumulating uniformly among neo-Y-linked genes, as expected if all loci on the neo-Y chromosome suffer from a reduced effectiveness of natural selection. In contrast, there is significant heterogeneity in the rate of protein evolution among neo-X-linked genes, with most loci being under strong purifying selection and two genes showing evidence for adaptive evolution. This observation agrees with theory predicting that linkage limits adaptive protein evolution. Both the neo-X and the neo-Y chromosome show an excess of unpreferred codon substitutions over preferred ones and no difference in this pattern was observed between the chromosomes. This suggests that there has been little or no selection maintaining codon bias in the D. miranda lineage. A change in mutational bias toward AT substitutions also contributes to the decline in codon bias. The contrast in patterns of molecular evolution between amino acid mutations and synonymous mutations on the neo-sex-linked genes can be understood in terms of chromosome-specific differences in effective population size and the distribution of selective effects of mutations.