Neutrality tests on mtDNA: unusual results from nematodes

McDonald-Kreitman tests of neutrality on mitochondrial DNA (mtDNA) of butterflies, Drosophila, and a variety of vertebrates usually show excess (over the neutral expectation) intraspecific polymorphism at nonsilent sites. These results are of great interest because they are the opposite of what is usually found for nuclear genes, in which the neutral pattern or evidence of adaptive divergence between species is usually observed. However, only vertebrates and insects have been tested so far, so it is not clear whether this intriguing pattern is typical for mtDNA in all taxa. Here I tested three pairs of nematode species and found that they all show a deficit of replacement polymorphism. Taken at face value, this result suggests that adaptive evolution proceeds more efficiently in nematode mtDNA than in mtDNA of vertebrates or insects. An alternate explanation is that the nematode pattern is an artifact of silent-site saturation that results from the rapid and composition-biased way in which nematode mtDNA evolves. Further studies are needed to distinguish between these two hypotheses.     

Mitonuclear Coevolution,but not Nuclear Compensation,Drives Evolution of OXPHOS Complexes in Bivalves

In Metazoa, four out of five complexes involved in oxidative phosphorylation (OXPHOS) are formed by subunits encoded by both the mitochondrial (mtDNA) and nuclear (nuDNA) genomes, leading to the expectation of mitonuclear coevolution. Previous studies have supported coadaptation of mitochondria-encoded (mtOXPHOS) and nuclear-encoded OXPHOS (nuOXPHOS) subunits, often specifically interpreted with regard to the “nuclear compensation hypothesis,” a specific form of mitonuclear coevolution where nuclear genes compensate for deleterious mitochondrial mutations due to less efficient mitochondrial selection. In this study, we analyzed patterns of sequence evolution of 79 OXPHOS subunits in 31 bivalve species, a taxon showing extraordinary mtDNA variability and including species with “doubly uniparental” mtDNA inheritance. Our data showed strong and clear signals of mitonuclear coevolution. NuOXPHOS subunits had concordant topologies with mtOXPHOS subunits, contrary to previous phylogenies based on nuclear genes lacking mt interactions. Evolutionary rates between mt and nuOXPHOS subunits were also highly correlated compared with non-OXPHO-interacting nuclear genes. Nuclear subunits of chimeric OXPHOS complexes (I, III, IV, and V) also had higher dN/dS ratios than Complex II, which is formed exclusively by nuDNA-encoded subunits. However, we did not find evidence of nuclear compensation: mitochondria-encoded subunits showed similar dN/dS ratios compared with nuclear-encoded subunits, contrary to most previously studied bilaterian animals. Moreover, no site-specific signals of compensatory positive selection were detected in nuOXPHOS genes. Our analyses extend the evidence for mitonuclear coevolution to a new taxonomic group, but we propose a reconsideration of the nuclear compensation hypothesis.     

Synonymous and nonsynonymous polymorphisms versus divergences in bacterial genomes

Comparison of the ratio of nonsynonymous to synonymous polymorphisms within species with the ratio of nonsynonymous to synonymous substitutions between species has been widely used as a supposed indicator of positive Darwinian selection, with the ratio of these 2 ratios being designated as a neutrality index (NI). Comparison of genome-wide polymorphism within 12 species of bacteria with divergence from an outgroup species showed substantial differences in NI among taxa. A low level of nonsynonymous polymorphism at a locus was the best predictor of NI < 1, rather than a high level of nonsynonymous substitution between species. Moreover, genes with NI < 1 showed a strong tendency toward the occurrence of rare nonsynonymous polymorphisms, as expected under the action of ongoing purifying selection. Thus, our results are more consistent with the hypothesis that a high relative rate of between-species nonsynonymous substitution reflects mainly the action of purifying selection within species to eliminate slightly deleterious mutations rather than positive selection between species. This conclusion is consistent with previous results highlighting an important role of slightly deleterious variants in bacterial evolution and suggests caution in the use of the McDonald-Kreitman test and related statistics as tests of positive selection.     

Positive selection in alternatively spliced exons of human genes

Alternative splicing is a well-recognized mechanism of accelerated genome evolution. We have studied single-nucleotide polymorphisms and human-chimpanzee divergence in the exons of 6672 alternatively spliced human genes, with the aim of understanding the forces driving the evolution of alternatively spliced sequences. Here, we show that alternatively spliced exons and exon fragments (alternative exons) from minor isoforms experience lower selective pressure at the amino acid level, accompanied by selection against synonymous sequence variation. The results of the McDonald-Kreitman test suggest that alternatively spliced exons, unlike exons constitutively included in the mRNA, are also subject to positive selection, with up to 27% of amino acids fixed by positive selection.     

Evolution of interacting proteins in the mitochondrial electron transport system in a marine copepod

The extensive interaction between mitochondrial-encoded and nuclear-encoded subunits of electron transport system (ETS) enzymes in mitochondria is expected to lead to intergenomic coadaptation. Whether this coadaptation results from adaptation to the environment or from fixation of deleterious mtDNA mutations followed by compensatory nuclear gene evolution is unknown. The intertidal copepod Tigriopus californicus shows extreme divergence in mtDNA sequence and provides an excellent model system for study of intergenomic coadaptation. Here, we examine genes encoding subunits of complex III of the ETS, including the mtDNA-encoded cytochrome b (CYTB), the nuclear-encoded rieske iron-sulfur protein (RISP), and cytochrome c(1) (CYC1). We compare levels of polymorphism within populations and divergence between populations in these genes to begin to untangle the selective forces that have shaped evolution in these genes. CYTB displays dramatic divergence between populations, but sequence analysis shows no evidence for positive selection driving this divergence. CYC1 and RISP have lower levels of sequence divergence between populations than CYTB, but, again, sequence analysis gives no evidence for positive selection acting on them. However, an examination of variation at cytochrome c (CYC), a nuclear-encoded protein that transfers electrons between complex III and complex IV provides evidence for selective divergence. Hence, it appears that rapid evolution in mitochondrial-encoded subunits is not always associated with rapid divergence in interacting subunits (CYC1 and RISP), but can be in some cases (CYC). Finally, a comparison of nuclear-encoded and mitochondrial-encoded genes from T. californicus suggests that substitution rates in the mitochondrial-encoded genes are dramatically increased relative to nuclear genes.     

MULTIPLE MODES OF SPECIATION INVOLVED IN THE PARALLEL EVOLUTION OF SYMPATRIC MORPHOTYPES OF LAKE WHITEFISH (COREGONUS CLUPEAFORMIS,SALMONIDAE)

We performed a phylogenetic analysis of mtDNA variation among seven sympatric pairs of dwarf and normal morphotypes of whitefish from northern Québec and the St. John River drainage to address three questions relevant to understanding their radiation. Are all sympatric pairs reproductively isolated? Do phylogenetic analyses confirm that sympatric whitefish morphotypes found in eastern North America represent the outcome of polyphyletic evolutionary events? If so, did all sympatric pairs from the St. John River drainage originate from the same scenario of allopatric divergence and secondary contact? The hypothesis of genetic differentiation was supported for all sympatric pairs from the St. John River drainage, whereas lack of mtDNA diversity precluded any test of reproductive isolation for northern Québec populations. Patterns of mtDNA variation confirmed that dwarf and normal morphotypes evolved in parallel among independent, yet closely related, lineages, thus providing indirect evidence for the role of natural selection in promoting phenotypic radiation in whitefish. Patterns of mtDNA diversity among sympatric pairs of the St. John River indicated a complex picture of whitefish evolution that implied sympatric divergence and multiple allopatric divergence/secondary contact events on a small geographic scale. These results suggests that ecological opportunities, namely trophic niche availability, may promote population divergence in whitefish.     

Between-host evolution of cytotoxic T-lymphocyte epitopes in human immunodeficiency virus type 1: an approach based on phylogenetically independent comparisons

In human immunodeficiency virus type 1 (HIV-1), mutations that escape from cytotoxic T-lymphocyte (CTL) recognition have been documented, and sequence analyses have provided indirect support for the hypothesis that natural selection has favored CTL escape mutants within an infected host. In spite of such evidence for within-host selection by CTL, it has been more difficult to determine how natural selection by host CTL has influenced long-term evolution of HIV-1. We used statistical analysis of published HIV-1 genomic sequences to examine the role of natural selection in between-host evolution of CTL epitopes. Based on a phylogenetic analysis, we identified 21 pairs of closely related genomes isolated from different hosts and examined the pattern of nucleotide substitution in genomic regions encoding well-characterized CTL epitopes. The results revealed that certain CTL epitopes have been subject to repeated positive selection across the population, while others are generally conserved. Furthermore, evidence of positive selection was associated with divergence from the canonical epitope sequence and with an enhanced frequency of convergent amino acid sequence changes in CTL epitopes. The results support the hypothesis that CTL-driven selection has been a major factor in the long-term evolution of HIV-1.     

Monophyly of clade III nematodes is not supported by phylogenetic analysis of complete mitochondrial genome sequences

Background

The orders Ascaridida, Oxyurida, and Spirurida represent major components of zooparasitic nematode diversity, including many species of veterinary and medical importance. Phylum-wide nematode phylogenetic hypotheses have mainly been based on nuclear rDNA sequences, but more recently complete mitochondrial (mtDNA) gene sequences have provided another source of molecular information to evaluate relationships. Although there is much agreement between nuclear rDNA and mtDNA phylogenies, relationships among certain major clades are different. In this study we report that mtDNA sequences do not support the monophyly of Ascaridida, Oxyurida and Spirurida (clade III) in contrast to results for nuclear rDNA. Results from mtDNA genomes show promise as an additional independently evolving genome for developing phylogenetic hypotheses for nematodes, although substantially increased taxon sampling is needed for enhanced comparative value with nuclear rDNA. Ultimately, topological incongruence (and congruence) between nuclear rDNA and mtDNA phylogenetic hypotheses will need to be tested relative to additional independent loci that provide appropriate levels of resolution.

Results

For this comparative phylogenetic study, we determined the complete mitochondrial genome sequences of three nematode species, Cucullanus robustus (13,972 bp) representing Ascaridida, Wellcomia siamensis (14,128 bp) representing Oxyurida, and Heliconema longissimum (13,610 bp) representing Spirurida. These new sequences were used along with 33 published nematode mitochondrial genomes to investigate phylogenetic relationships among chromadorean orders. Phylogenetic analyses of both nucleotide and amino acid sequence datasets support the hypothesis that Ascaridida is nested within Rhabditida. The position of Oxyurida within Chromadorea varies among analyses; in most analyses this order is sister to the Ascaridida plus Rhabditida clade, with representative Spirurida forming a distinct clade, however, in one case Oxyurida is sister to Spirurida. Ascaridida, Oxyurida, and Spirurida (the sampled clade III taxa) do not form a monophyletic group based on complete mitochondrial DNA sequences. Tree topology tests revealed that constraining clade III taxa to be monophyletic, given the mtDNA datasets analyzed, was a significantly worse result.

Conclusion

The phylogenetic hypotheses from comparative analysis of the complete mitochondrial genome data (analysis of nucleotide and amino acid datasets, and nucleotide data excluding 3^rd positions) indicates that nematodes representing Ascaridida, Oxyurida and Spirurida do not share an exclusive most recent common ancestor, in contrast to published results based on nuclear ribosomal DNA. Overall, mtDNA genome data provides reliable support for nematode relationships that often corroborates findings based on nuclear rDNA. It is anticipated that additional taxonomic sampling will provide a wealth of information on mitochondrial genome evolution and sequence data for developing phylogenetic hypotheses for the phylum Nematoda.

     

Dntf-2r,a young Drosophila retroposed gene with specific male expression under positive Darwinian selection

A direct approach to investigating new gene origination is to examine recently evolved genes. We report a new gene in the Drosophila melanogaster subgroup, Drosophila nuclear transport factor-2-related (Dntf-2r). Its sequence features and phylogenetic distribution indicate that Dntf-2r is a retroposed functional gene and originated in the common ancestor of D. melanogaster, D. simulans, D. sechellia, and D. mauritiana, within the past 3-12 million years (MY). Dntf-2r evolved more rapidly than the parental gene, under positive Darwinian selection as revealed by the McDonald-Kreitman test and other evolutionary analyses. Comparative expression analysis shows that Dntf-2r is male specific whereas the parental gene, Dntf-2, is widely expressed in D. melanogaster. In agreement with its new expression pattern, the Dntf-2r putative promoter sequence is similar to the late testis promoter of beta2-tubulin. We discuss the possibility that the action of positive selection in Dntf-2r is related to its putative male-specific functions.     

Excess amino acid polymorphism in mitochondrial DNA: contrasts among genes from Drosophila, mice, and humans

Recent studies of mitochondrial DNA (mtDNA) variation in mammals and Drosophila have shown an excess of amino acid variation within species (replacement polymorphism) relative to the number of silent and replacement differences fixed between species. To examine further this pattern of nonneutral mtDNA evolution, we present sequence data for the ND3 and ND5 genes from 59 lines of Drosophila melanogaster and 29 lines of D. simulans. Of interest are the frequency spectra of silent and replacement polymorphisms, and potential variation among genes and taxa in the departures from neutral expectations. The Drosophila ND3 and ND5 data show no significant excess of replacement polymorphism using the McDonald-Kreitman test. These data are in contrast to significant departures from neutrality for the ND3 gene in mammals and other genes in Drosophila mtDNA (cytochrome b and ATPase 6). Pooled across genes, however, both Drosophila and human mtDNA show very significant excesses of amino acid polymorphism. Silent polymorphisms at ND5 show a significantly higher variance in frequency than replacement polymorphisms, and the latter show a significant skew toward low frequencies (Tajima's D = -1.954). These patterns are interpreted in light of the nearly neutral theory where mildly deleterious amino acid haplotypes are observed as ephemeral variants within species but do not contribute to divergence. The patterns of polymorphism and divergence at charge-altering amino acid sites are presented for the Drosophila ND5 gene to examine the evolution of functionally distinct mutations. Excess charge-altering polymorphism is observed at the carboxyl terminal and excess charge-altering divergence is detected at the amino terminal. While the mildly deleterious model fits as a net effect in the evolution of nonrecombining mitochondrial genomes, these data suggest that opposing evolutionary pressures may act on different regions of mitochondrial genes and genomes.      

Evolutionary Processes Acting on Candidate cis-Regulatory Regions in Humans Inferred from Patterns of Polymorphism and Divergence

Analysis of polymorphism and divergence in the non-coding portion of the human genome yields crucial information about factors driving the evolution of gene regulation. Candidate cis-regulatory regions spanning more than 15,000 genes in 15 African Americans and 20 European Americans were re-sequenced and aligned to the chimpanzee genome in order to identify potentially functional polymorphism and to characterize and quantify departures from neutral evolution. Distortions of the site frequency spectra suggest a general pattern of selective constraint on conserved non-coding sites in the flanking regions of genes (CNCs). Moreover, there is an excess of fixed differences that cannot be explained by a Gamma model of deleterious fitness effects, suggesting the presence of positive selection on CNCs. Extensions of the McDonald-Kreitman test identified candidate cis-regulatory regions with high probabilities of positive and negative selection near many known human genes, the biological characteristics of which exhibit genome-wide trends that differ from patterns observed in protein-coding regions. Notably, there is a higher probability of positive selection in candidate cis-regulatory regions near genes expressed in the fetal brain, suggesting that a larger portion of adaptive regulatory changes has occurred in genes expressed during brain development. Overall we find that natural selection has played an important role in the evolution of candidate cis-regulatory regions throughout hominid evolution.     

Nuclear markers confirm taxonomic status and relationships among highly endangered and closely related right whale species

Right whales (genus: Eubalaena) are among the most endangered mammals, yet their taxonomy and phylogeny have been questioned. A phylogenetic hypothesis based on mitochondrial DNA (mtDNA) variation recently prompted a taxonomic revision, increasing the number of right whale species to three. We critically evaluated this hypothesis using sequence data from 13 nuclear DNA (nuDNA) loci as well as the mtDNA control region. Fixed diagnostic characters among the nuclear markers strongly support the hypothesis of three genetically distinct species, despite lack of any diagnostic morphological characters. A phylogenetics analysis of all data produced a strict consensus cladogram with strong support at nodes that define each right whale species as well as relationships among species. Results showed very little conflict among the individual partitions as well as congruence between the mtDNA and nuDNA datasets. These data clearly demonstrate the strength of using numerous independent genetic markers during a phylogenetics analysis of closely related species. In evaluating phylogenetic support contributed by individual loci, 11 of the 14 loci provided support for at least one of the nodes of interest to this study. Only a single marker (mtDNA control region) provided support at all four nodes. A study using any single nuclear marker would have failed to support the proposed phylogeny, and a strong phylogenetic hypothesis was only revealed by the simultaneous analysis of many nuclear loci. In addition, nu DNA and mtDNA data provided complementary levels of support at nodes of different evolutionary depth indicating that the combined use of mtDNA and nuDNA data is both practical and desirable.     

Weighing the evidence for adaptation at the molecular level

The abundance of genome polymorphism and divergence data has provided unprecedented insight into how mutation, drift and natural selection shape genome evolution. Application of the McDonald-Kreitman (MK) test to such data indicates a pervasive influence of positive selection, particularly in Drosophila species. However, evidence for positive selection in other species ranging from yeast to humans is often weak or absent. Although evidence for positive selection could be obscured in some species, there is also reason to believe that the frequency of adaptive substitutions could be overestimated as a result of epistatic fitness effects or hitchhiking of deleterious mutations. Based on these considerations it is argued that the common assumption of independence among sites must be relaxed before abandoning the neutral theory of molecular evolution.     

Multiple nuclear gene sequences identify phylogenetic species boundaries in the rapidly radiating clade of Mexican ambystomatid salamanders

Delimiting the boundaries of species involved in radiations is critical to understanding the tempo and mode of lineage formation. Single locus gene trees may or may not reflect the underlying pattern of population divergence and lineage formation, yet they constitute the vast majority of the empirical data in species radiations. In this study we make use of an expressed sequence tag (EST) database to perform nuclear (nDNA) and mitochondrial (mtDNA) genealogical tests of species boundaries in Ambystoma ordinarium, a member of an adaptive radiation of metamorphic and paedomorphic salamanders (the Ambystoma tigrinum complex) that have diversified across terrestrial and aquatic environments. Gene tree comparisons demonstrate extensive nonmonophyly in the mtDNA genealogy of A. ordinarium, while seven of eight independent nuclear loci resolve the species as monophyletic or nearly so, and diagnose it as a well-resolved genealogical species. A differential introgression hypothesis is supported by the observation that western A. ordinarium localities contain mtDNA haplotypes that are identical or minimally diverged from haplotypes sampled from a nearby paedomorphic species, Ambystoma dumerilii, while most nDNA trees place these species in distant phylogenetic positions. These results provide a strong example of how historical introgression can lead to radical differences between gene trees and species histories, even among currently allopatric species with divergent life history adaptations and morphologies. They also demonstrate how EST-based nuclear resources can be used to more fully resolve the phylogenetic history of species radiations.     

Parallel evolution and vicariance in the guppy (Poecilia reticulata) over multiple spatial and temporal scales

Well-studied model systems present ideal opportunities to understand the relative roles of contemporary selection versus historical processes in determining population differentiation and speciation. Although guppy populations in Trinidad have been a model for studies of evolutionary ecology and sexual selection for more than 50 years, this work has been conducted with little understanding of the phylogenetic history of this species. We used variation in nuclear (X-src) and mitochondrial DNA (mtDNA) sequences to examine the phylogeographic history of Poecilia reticulata Peters (the guppy) across its entire natural range, and to test whether patterns of morphological divergence are a consequence of parallel evolution. Phylogenetic, nested clade, population genetic, and demographic analyses were conducted to investigate patterns of genetic structure at several temporal scales and are discussed in relation to vicariant events, such as tectonic activity and glacial cycles, shaping northeast South American river drainages. The mtDNA phylogeny defined five major lineages, each associated with one or more river drainages, and analysis of molecular variance also detected geographic structuring among these river drainages in an evolutionarily conserved nuclear (X-src) locus. Nested clade and other demographic analyses suggest that the eastern Venezuela/ western Trinidad region is likely the center of origin of P. reticulata. Mantel tests show that the divergence of morphological characters, known to differentiate on a local scale in response to natural and sexual selection pressures, is not associated with mtDNA genetic distance; however, TreeScan analysis identified several significant associations of these characters with the haplotype tree. Parallel upstream/downstream patterns of morphological adaptation in response to selection pressures reported in P. reticulata within Trinidad rivers appears to persist across the natural range. Our results together with previous studies suggest that, although morphological variation in P. reticulata is primarily attributed to selection, phylogeographic history may also play a role.     

Phylogenetic codivergence supports coevolution of mimetic Heliconius butterflies

The unpalatable and warning-patterned butterflies Heliconius erato and Heliconius melpomene provide the best studied example of mutualistic Müllerian mimicry, thought-but rarely demonstrated-to promote coevolution. Some of the strongest available evidence for coevolution comes from phylogenetic codivergence, the parallel divergence of ecologically associated lineages. Early evolutionary reconstructions suggested codivergence between mimetic populations of H. erato and H. melpomene, and this was initially hailed as one of the most striking known cases of coevolution. However, subsequent molecular phylogenetic analyses found discrepancies in phylogenetic branching patterns and timing (topological and temporal incongruence) that argued against codivergence. We present the first explicit cophylogenetic test of codivergence between mimetic populations of H. erato and H. melpomene, and re-examine the timing of these radiations. We find statistically significant topological congruence between multilocus coalescent population phylogenies of H. erato and H. melpomene. Cophylogenetic historical reconstructions support repeated codivergence of mimetic populations, from the base of the sampled radiations. Pairwise distance correlation tests, based on our coalescent analyses plus recently published AFLP and wing colour pattern gene data, also suggest that the phylogenies of H. erato and H. melpomene show significant topological congruence. Divergence time estimates, based on a Bayesian coalescent model, suggest that the evolutionary radiations of H. erato and H. melpomene occurred over the same time period, and are compatible with a series of temporally congruent codivergence events. Our results suggest that differences in within-species genetic divergence are the result of a greater overall effective population size for H. erato relative to H. melpomene and do not imply incongruence in the timing of their phylogenetic radiations. Repeated codivergence between Müllerian co-mimics, predicted to exert mutual selection pressures, strongly suggests coevolution. Our results therefore support a history of reciprocal coevolution between Müllerian co-mimics characterised by phylogenetic codivergence and parallel phenotypic change.     

Reproductive protein evolution in two cryptic species of marine chordate

Background  

Reproductive character displacement (RCD) is a common and taxonomically widespread pattern. In marine broadcast spawning organisms, behavioral and mechanical isolation are absent and prezygotic barriers between species often operate only during the fertilization process. Such barriers are usually a consequence of differences in the way in which sperm and egg proteins interact, so RCD can be manifest as faster evolution of these proteins between species in sympatry than allopatry. Rapid evolution of these proteins often appears to be a consequence of positive (directional) selection. Here, we identify a set of candidate gamete recognition proteins (GRPs) in the ascidian Ciona intestinalis and showed that these GRPs evolve more rapidly than control proteins (those not involved in gamete recognition). Choosing a subset of these gamete recognition proteins that show evidence of positive selection (CIPRO37.40.1, CIPRO60.5.1, CIPRO100.7.1), we then directly test the RCD hypothesis by comparing divergence (omega) and polymorphism (McDonald-Kreitman, Tajima's D, Fu and Li's D and F, Fay and Wu's H) statistics in sympatric and allopatric populations of two distinct forms of C. intestinalis (Types A and B) between which there are strong post-zygotic barriers.     

An investigation of the statistical power of neutrality tests based on comparative and population genetic data

In this report, we investigate the statistical power of several tests of selective neutrality based on patterns of genetic diversity within and between species. The goal is to compare tests based solely on population genetic data with tests using comparative data or a combination of comparative and population genetic data. We show that in the presence of repeated selective sweeps on relatively neutral background, tests based on the d(N)/d(S) ratios in comparative data almost always have more power to detect selection than tests based on population genetic data, even if the overall level of divergence is low. Tests based solely on the distribution of allele frequencies or the site frequency spectrum, such as the Ewens-Watterson test or Tajima's D, have less power in detecting both positive and negative selection because of the transient nature of positive selection and the weak signal left by negative selection. The Hudson-Kreitman-Aguadé test is the most powerful test for detecting positive selection among the population genetic tests investigated, whereas McDonald-Kreitman test typically has more power to detect negative selection. We discuss our findings in the light of the discordant results obtained in several recently published genomic scans.     

Evolutionary significance of gene expression divergence

Recent large-scale studies of evolutionary changes in gene expression among mammalian species have led to the proposal that gene expression divergence may be neutral with respect to organismic fitness. Here, we employ a comparative analysis of mammalian gene sequence divergence and gene expression divergence to test the hypothesis that the evolution of gene expression is predominantly neutral. Two models of neutral gene expression evolution are considered: 1-purely neutral evolution (i.e., no selective constraint) of gene expression levels and patterns and 2-neutral evolution accompanied by selective constraint. With respect to purely neutral evolution, levels of change in gene expression between human-mouse orthologs are correlated with levels of gene sequence divergence that are determined largely by purifying selection. In contrast, evolutionary changes of tissue-specific gene expression profiles do not show such a correlation with sequence divergence. However, divergence of both gene expression levels and profiles are significantly lower for orthologous human-mouse gene pairs than for pairs of randomly chosen human and mouse genes. These data clearly point to the action of selective constraint on gene expression divergence and are inconsistent with the purely neutral model; however, there is likely to be a neutral component in evolution of gene expression, particularly, in tissues where the expression of a given gene is low and functionally irrelevant. The model of neutral evolution with selective constraint predicts a regular, clock-like accumulation of gene expression divergence. However, relative rate tests of the divergence among human-mouse-rat orthologous gene sets reveal clock-like evolution for gene sequence divergence, and to a lesser extent for gene expression level divergence, but not for the divergence of tissue-specific gene expression profiles. Taken together, these results indicate that gene expression divergence is subject to the effects of purifying selective constraint and suggest that it might also be substantially influenced by positive Darwinian selection.     

Widespread adaptive evolution of Drosophila genes with sex-biased expression

Many genes in higher eukaryotes show sexually dimorphic expression, and these genes tend to be among the most divergent between species. In most cases, however, it is not known whether this rapid divergence is caused by positive selection or if it is due to a relaxation of selective constraint. To distinguish between these two possibilities, we surveyed DNA sequence polymorphism in 91 Drosophila melanogaster genes with male-, female-, or nonsex-biased expression and determined their divergence from the sister species D. simulans. Using several single- and multilocus statistical tests, we estimated the type and strength of selection influencing the evolution of the proteins encoded by genes of each expression class. Adaptive evolution, as indicated by a relative excess of nonsynonymous divergence between species, was common among the sex-biased genes (both male and female). Male-biased genes, in particular, showed a strong and consistent signal of positive selection, while female-biased genes showed more variation in the type of selection they experience. Genes expressed equally in the two sexes, in contrast, showed no evidence for adaptive evolution between D. melanogaster and D. simulans. This suggests that sexual selection and intersexual coevolution are the major forces driving genetic differentiation between species.