Evidence of adaptive evolution of accessory gland proteins in closely related species of the Drosophila repleta group

Accessory gland proteins (Acps) are part of the seminal fluid of Drosophila species. These proteins have important reproductive functions, being responsible for the proper functioning of several steps of the fertilization process. Acps also contribute indirectly for the reproductive success of males by modulating female behavior. Evidence that Acps participate in sperm competition and sexual conflict includes findings that, on average, Acps have fast evolutionary rates, suggestive of adaptive evolution. This is especially true in species of the Drosophila repleta group. Nevertheless, only in a few occasions have robust statistical tests been used to determine whether observed evolutionary rates are in fact due to positive selection on amino acid substitutions between related species. Here we apply maximum likelihood tests for positive selection on 14 Acps of the D. repleta group. To increase statistical robustness, we use at least 8 sequences, all belonging to species of the Drosophila mulleri complex, for each gene analyzed. We found significant evidence of adaptive evolution for 10 of the tested genes. Among these, the ones with a conserved protein domain had positively selected sites within the functional region of the sequence. We also detected one instance of lineage-specific adaptive evolution in a clade formed by 2 sister species.     

The role of GC-biased gene conversion in shaping the fastest evolving regions of the human genome

GC-biased gene conversion (gBGC) is a recombination-associated evolutionary process that accelerates the fixation of guanine or cytosine alleles, regardless of their effects on fitness. gBGC can increase the overall rate of substitutions, a hallmark of positive selection. Many fast-evolving genes and noncoding sequences in the human genome have GC-biased substitution patterns, suggesting that gBGC-in contrast to adaptive processes-may have driven the human changes in these sequences. To investigate this hypothesis, we developed a substitution model for DNA sequence evolution that quantifies the nonlinear interacting effects of selection and gBGC on substitution rates and patterns. Based on this model, we used a series of lineage-specific likelihood ratio tests to evaluate sequence alignments for evidence of changes in mode of selection, action of gBGC, or both. With a false positive rate of less than 5% for individual tests, we found that the majority (76%) of previously identified human accelerated regions are best explained without gBGC, whereas a substantial minority (19%) are best explained by the action of gBGC alone. Further, more than half (55%) have substitution rates that significantly exceed local estimates of the neutral rate, suggesting that these regions may have been shaped by positive selection rather than by relaxation of constraint. By distinguishing the effects of gBGC, relaxation of constraint, and positive selection we provide an integrated analysis of the evolutionary forces that shaped the fastest evolving regions of the human genome, which facilitates the design of targeted functional studies of adaptation in humans.     

Adaptive evolution of genes underlying schizophrenia

Schizophrenia poses an evolutionary-genetic paradox because it exhibits strongly negative fitness effects and high heritability, yet it persists at a prevalence of approximately 1% across all human cultures. Recent theory has proposed a resolution: that genetic liability to schizophrenia has evolved as a secondary consequence of selection for human cognitive traits. This hypothesis predicts that genes increasing the risk of this disorder have been subject to positive selection in the evolutionary history of humans and other primates. We evaluated this prediction using tests for recent selective sweeps in human populations and maximum-likelihood tests for selection during primate evolution. Significant evidence for positive selection was evident using one or both methods for 28 of 76 genes demonstrated to mediate liability to schizophrenia, including DISC1, DTNBP1 and NRG1, which exhibit especially strong and well-replicated functional and genetic links to this disorder. Strong evidence of non-neutral, accelerated evolution was found for DISC1, particularly for exon 2, the only coding region within the schizophrenia-associated haplotype. Additionally, genes associated with schizophrenia exhibited a statistically significant enrichment in their signals of positive selection in HapMap and PAML analyses of evolution along the human lineage, when compared with a control set of genes involved in neuronal activities. The selective forces underlying adaptive evolution of these genes remain largely unknown, but these findings provide convergent evidence consistent with the hypothesis that schizophrenia represents, in part, a maladaptive by-product of adaptive changes during human evolution.     

A scan for signatures of positive selection in candidate loci for skin pigmentation in humans

Although the combination of pale skin and intense sun exposure results in an important health risk for the individual, it is less clear if at the population level this risk has possessed an evolutionary meaning. In this sense, a number of adaptive hypotheses have been put forward to explain the evolution of human skin pigmentation, such as photoprotection against sun-induced cancer, sexual selection, vitamin D synthesis or photoprotection of photolabile compounds, among others. It is expected that if skin pigmentation is adaptive, we might be able to see the signature of positive selection on some of the genes involved. In order to detect this signature, we analyze a battery of 81 candidate loci by means of phylogenetic and population genetic tests. Our results indicate that both light and dark skin may possess adaptive value. Of the main loci presenting this signature, TP53BP1 shows clear evidence of adaptive selection in Africans, whereas TYRP1 and SLC24A5 show evidence of adaptive selection in Caucasians. Although we cannot offer a mechanism that based on these genes explains the advantage of light skin, if TP53BP1, and perhaps RAD50, have truly conferred an adaptive value to the African population analyzed, photoprotection against sun-induced skin damage/cancer might be proposed as a mechanism that has driven the evolution of human skin pigmentation.     

Phycoerythrin evolution and diversification of spectral phenotype in marine Synechococcus and related picocyanobacteria

In marine Synechococcus there is evidence for the adaptive evolution of spectrally distinct forms of the major light harvesting pigment phycoerythrin (PE). Recent research has suggested that these spectral forms of PE have a different evolutionary history than the core genome. However, a lack of explicit statistical testing of alternative hypotheses or for selection on these genes has made it difficult to evaluate the evolutionary relationships between spectral forms of PE or the role horizontal gene transfer (HGT) may have had in the adaptive phenotypic evolution of the pigment system in marine Synechococcus. In this work, PE phylogenies of picocyanobacteria with known spectral phenotypes, including newly co-isolated strains of marine Synechococcus from the Gulf of Mexico, were constructed to explore the diversification of spectral phenotype and PE evolution in this group more completely. For the first time, statistical evaluation of competing evolutionary hypotheses and tests for positive selection on the PE locus in picocyanobacteria were performed. Genes for PEs associated with specific PE spectral phenotypes formed strongly supported monophyletic clades within the PE tree with positive directional selection driving evolution towards higher phycourobilin (PUB) content. The presence of the PUB-lacking phenotype in PE-containing marine picocyanobacteria from cyanobacterial lineages identified as Cyanobium is best explained by HGT into this group from marine Synechococcus. Taken together, these data provide strong examples of adaptive evolution of a single phenotypic trait in bacteria via mutation, positive directional selection and horizontal gene transfer.     

Positive selection in East Asians for an EDAR allele that enhances NF-kappaB activation

Genome-wide scans for positive selection in humans provide a promising approach to establish links between genetic variants and adaptive phenotypes. From this approach, lists of hundreds of candidate genomic regions for positive selection have been assembled. These candidate regions are expected to contain variants that contribute to adaptive phenotypes, but few of these regions have been associated with phenotypic effects. Here we present evidence that a derived nonsynonymous substitution (370A) in EDAR, a gene involved in ectodermal development, was driven to high frequency in East Asia by positive selection prior to 10,000 years ago. With an in vitro transfection assay, we demonstrate that 370A enhances NF-kappaB activity. Our results suggest that 370A is a positively selected functional genetic variant that underlies an adaptive human phenotype.     

Positive Selection in East Asians for an EDAR Allele that Enhances NF-κB Activation

Genome-wide scans for positive selection in humans provide a promising approach to establish links between genetic variants and adaptive phenotypes. From this approach, lists of hundreds of candidate genomic regions for positive selection have been assembled. These candidate regions are expected to contain variants that contribute to adaptive phenotypes, but few of these regions have been associated with phenotypic effects. Here we present evidence that a derived nonsynonymous substitution (370A) in EDAR, a gene involved in ectodermal development, was driven to high frequency in East Asia by positive selection prior to 10,000 years ago. With an in vitro transfection assay, we demonstrate that 370A enhances NF-κB activity. Our results suggest that 370A is a positively selected functional genetic variant that underlies an adaptive human phenotype.     

A population genomic approach to map recent positive selection in model species

Based on nearly complete genome sequences from a variety of organisms data on naturally occurring genetic variation on the scale of hundreds of loci to entire genomes have been collected in recent years. In parallel, new statistical tests have been developed to infer evidence of recent positive selection from these data and to localize the target regions of selection in the genome. These methods have now been successfully applied to Drosophila melanogaster , humans, mice and a few plant species. In genomic regions of normal recombination rates, the targets of positive selection have been mapped down to the level of individual genes.     

Compound tests for the detection of hitchhiking under positive selection

Many statistical tests have been developed for detecting positive selection. Most of these tests draw conclusions based on significant deviations from the patterns of polymorphism predicted by the neutral model. However, many non-equilibrium forces may cause similar deviations, and thus the tests usually have low statistical specificity to positive selection. The main challenge is hence to construct test statistics that are reasonably powerful in detecting positive selection, but are relatively insensitive to other forces. Recently, Zeng et al. (2006) proposed a new test, DH, which is a compound of Tajima's D and Fay and Wu's H, and showed that DH has reasonably high statistical specificity to positive selection. In this report, we expand the idea of a compound test by combining Fay and Wu's H or DH with the Ewens-Watterson (EW) test. We refer to these 2 new tests as HEW and DHEW, respectively. Compared to the DH test, HEW and DHEW are more robust against the presence of recombination, and are also more powerful in detecting positive selection. Furthermore, the DHEW test, similar to DH, is also relatively insensitive to background selection and demography. The HEW test, on the other hand, tends to be somewhat less conservative than DH and DHEW in some cases.     

A reassessment of explanations for discordant introgressions of mitochondrial and nuclear genomes

Hybridization is increasingly recognized as a significant evolutionary process, in particular because it can lead to introgression of genes from one species to another. A striking pattern of discordance in the amount of introgression between mitochondrial and nuclear markers exists such that substantial mitochondrial introgression is often found in combination with no or little nuclear introgression. Multiple mechanisms have been proposed to explain this discordance, including positive selection for introgressing mitochondrial variants, several types of sex‐biases, drift, negative selection against introgression in the nuclear genome, and spatial expansion. Most of these hypotheses are verbal, and have not been quantitatively evaluated so far. We use individual‐based, multilocus, computer simulations of secondary contact under a wide range of demographic and genetic scenarios to evaluate the ability of the different mechanisms to produce discordant introgression. Sex‐biases and spatial expansions fail to produce substantial mito‐nuclear discordance. Drift and nuclear selection can produce strong discordance, but only under a limited range of conditions. In contrast, selection on the mitochondrial genome produces strong discordance, particularly when dispersal rates are low. However, commonly used statistical tests have little power to detect this selection. Altogether, these results dismiss several popular hypotheses, and provide support for adaptive mitochondrial introgression.     

Molecular diversity and evolution of myticin-C antimicrobial peptide variants in the Mediterranean mussel, Mytilus galloprovincialis

Mussels have diverse groups of cysteine rich, cationic antimicrobial peptides (AMPs) (defensins, mytilins, myticins, and mytimycin) that constitute an important component of their innate immune defence. Despite the identification and characterization of these AMPs in mussels, the underlying genetic mechanisms that maintain high diversity among multiple variants of the myticin-C isoform are poorly understood. Using phylogeny-based models of sequence evolution and several site-by-site frequency spectrum statistical tests for neutrality, herein we report that positive selection has been the major driving force in maintaining high diversity among the allelic-variants of the myticin-C AMP of Mytilus galloprovincialis. The statistical tests rejected the hypothesis that all polymorphism within myticin-C loci is neutral. Although a majority of the codons constrained to purifying selection (rate of amino acid replacement to the silent substitution, omega < 1), approximately 8% of the codons with omega approximately equal to 5.5 are under positive selection (omega > 1), thus indicating adaptive evolution of certain amino acids. Direct interaction of these peptides with the surrounding pathogens and/or altered/new pathogens in the changing environment is the likely cause of molecular adaptation of certain amino acid sites in myticin-C variants.     

Patterns of selection and polymorphism of innate immunity genes in bumblebees (Hymenoptera: Apidae)

In response to on-going biodiversity loss, conservation genetics has established itself as an important branch of biology. Historically concentrating on assessing stochastic processes using neutral loci, there has been a recent surge of interest in understanding and quantifying variation at loci underlying ecologically important traits. To this end, patterns of selection and polymorphism at these loci must be characterized. Loci underlying immunity make good candidates in this context: they are expected to be important for population persistence and may exhibit diversifying or divergent selection. Predictions regarding the pattern of selection expected at immune system loci have been based on their interactions with pathogens, however, published studies report mixed results as to whether these are borne out or not. Here, polymorphism and selection is examined for three innate immune system loci in bumblebees: a peptidoglycan recognition protein, a putative alpha-macroglobulin, and scavenger receptor. Both intra- and inter-specific sequence variation is quantified. Very little polymorphism was encountered, precluding robust tests of selection. However, the lack of inter-specific polymorphisms suggests a lack of positive selection for the regions sequenced. Results are discussed with respect to population genetic predictions and generation of a specific immune response in insects. Alternative loci and methods for studying adaptive genetic variation in a conservation context are considered.     

Adaptive evolution after duplication of penaeidin antimicrobial peptides

Penaeidin antimicrobial peptides in penaeid shrimps are an important component of their innate immune system that provides immunity against infection caused by several gram-positive bacteria and filamentous fungal species. Despite the knowledge on the identification and characterization of these peptides in penaeid shrimps, little is known about the evolutionary pattern of these peptides and the underlying genetic mechanisms that maintain high sequence diversities in the penaeidin gene family. Based on the phylogenetic analyses and maximum likelihood-based codon substitution analyses, here we present the convincing evidence that multiple copies of penaeidins have evolved by gene duplication, and positive Darwinian selection (adaptive evolution) is the likely cause of accelerated rate of amino acid substitutions among these duplicated genes. While the average ratio of non-synonymous to synonymous substitutions (omega) for the entire coding region of both active domains is 0.9805, few codon sites showed significantly higher omega (3.73). The likelihood ratio tests that compare models incorporating positive selection (omega>1) at certain codon sites with models not incorporating positive selection (omega<1), failed to reject (p=0) the evidence of positive Darwinian selection. The rapid adaptive evolution of this gene family might be directed by the pathogens and the faster rate of amino acid substitutions in the N-terminal proline-rich and C-terminal cysteine-rich domains could be due to their direct involvement in the protection against pathogens. When the host expose to different habitats/environment an accelerated rate of amino acid substitutions in both the active domains may also be expected.     

Complex multivariate sexual selection on male acoustic signaling in a wild population of Teleogryllus commodus

Mate choice may impose both linear (i.e., directional) and nonlinear (i.e., quadratic and correlational) sexual selection on advertisement traits. Traditionally, mate recognition and sensory tuning have been thought to impose stabilizing (i.e., negative quadratic) sexual selection, whereas adaptive mate choice effects directional selection. It has been suggested that adaptive choice may exert positive quadratic and/or correlational sexual selection. Earlier, we showed that five structural components of the advertisement call of male field crickets (Teleogryllus commodus) were under multivariate stabilizing selection under laboratory conditions. Here we experimentally estimate selection on these five traits plus a measure of calling activity (the number of repeats in a looped bout of calling) in the field. There was general support for multivariate stabilizing selection on call structure, and calling activity was under strong positive directional selection, as predicted for a signal of genetic quality. There was, however, also appreciable correlational selection, suggesting an interaction between male call structure and calling effort. Interestingly, selection for short interbout durations of silence favored longer intercall durations in the field, in contrast to results from continuous looped call playback in the laboratory. We discuss the general importance of nonlinear selection in the honest signaling of genetic quality.     

The influence of phylogenetic uncertainty on the detection of positive Darwinian selection

The power of maximum likelihood tests of positive selection on protein-coding genes depends heavily on detecting and accounting for potential biases in the studied data set. Although the influence of transition:transversion and codon biases have been investigated in detail, little is known about how inaccuracy in the phylogeny used during the calculations affects the performance of these tests. In this study, 3 empirical data sets are analyzed using sets of simulated topologies corresponding to low, intermediate, and high levels of phylogenetic uncertainty. The detection of positive selection was largely unaffected by errors in the underlying phylogeny. However, the number of sites identified as being under positive selection tended to be overestimated.     

Detecting strong positive selection in the genome

New statistical tests have been developed in the past decade that enable us to infer evidence of recent strong positive selection from genome-wide data on single-nucleotide polymorphism and to localize the targets of selection in the genome. Based on these tests, past demographic events that led to distortions of the site-frequency spectrum of variation can be distinguished from selection, in particular if linkage disequilibrium is taken into account. These methods have been successfully applied to species from which complete sequence information and polymorphism data are available, including Drosophila melanogaster, humans, and several plant species. To make full use of the available data, however, the tests that were primarily designed for panmictic populations need to be extended to spatially structured populations.     

Identifying genes underlying skin pigmentation differences among human populations

Skin pigmentation is a human phenotype that varies greatly among human populations and it has long been speculated that this variation is adaptive. We therefore expect the genes that contribute to these large differences in phenotype to show large allele frequency differences among populations and to possibly harbor signatures of positive selection. To identify the loci that likely contribute to among-population human skin pigmentation differences, we measured allele frequency differentiation among Europeans, Chinese and Africans for 24 human pigmentation genes from 2 publicly available, large scale SNP data sets. Several skin pigmentation genes show unusually large allele frequency differences among these populations. To determine whether these allele frequency differences might be due to selection, we employed a within-population test based on long-range haplotype structure and identified several outliers that have not been previously identified as putatively adaptive. Most notably, we identify the DCT gene as a candidate for recent positive selection in the Chinese. Moreover, our analyses suggest that it is likely that different genes are responsible for the lighter skin pigmentation found in different non-African populations. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Evidence of positively selected sites in mammalian alpha-defensins

Alpha-defensins are a family of mammalian antimicrobial peptides that exhibit variable activity against a panel of microbes, including bacteria, fungi, and enveloped viruses. We have employed a maximum-likelihood approach to detect evidence of positive selection (adaptive evolution) in the evolution of these important molecules of the innate immune response. We have identified 14 amino acid sites that are predicted to be subject to positive selection. Furthermore, we show that all these sites are located in the mature antimicrobial peptide and not in the prepropeptide region of the molecule, implying that they are of functional importance. These results suggest that mammalian alpha-defensins have been under selective pressure to evolve in response to potentially infectious challenges by fast-evolving microbes.     

An algorithm for detecting directional and non-directional positive selection,neutrality and negative selection in protein coding DNA sequences

Positive selection or adaptive evolution is thought to be responsible, at least some of the time, for the rapid accumulation of advantageous changes in protein-coding genes. The origin of new enzymatic functions, erection of barriers to heterospecific fertilization, and evasion of host response by pathogens, among other things, are thought to be instances of adaptive evolution. Detecting positive selection in protein-coding genes is fraught with difficulties. Saturation for sequence change, codon usage bias, ephemeral selection events and differential selective pressures on amino acids all contribute to the problem. A number of solutions have been proposed with varying degrees of success, however they suffer from limitations of not being accurate enough or being prohibitively computationally intensive. We have developed a character-based method of identifying lineages that undergo positive selection. In our method we assess the possibility that for each internal branch of a phylogenetic tree an event occurred that subsequently gave rise to a greater number of replacement substitutions than might be expected. We classify these replacement substitutions into two categories – whether they subsequently became invariable or changed again in at least one descendent lineage. The former situation indicates that the new character state is under strong selection to preserve its new identity (directional selection), while the latter situation indicates that there is a persistent pressure to change identity (non-directional selection). The method is fast and accurate, easy to implement, sensitive to short-lived selection events and robust with respect to sampling density and proportion of sites under the influence of positive selection.     

Molecular spandrels: tests of adaptation at the genetic level

Although much progress has been made in identifying the genes (and, in rare cases, mutations) that contribute to phenotypic variation, less is known about the effects that these genes have on fitness. Nonetheless, genes are commonly labelled as 'adaptive' if an allele has been shown to affect a phenotype with known or suspected functional importance or if patterns of nucleotide variation at the locus are consistent with positive selection. In these cases, the 'adaptive' designation may be premature and may lead to incorrect conclusions about the relationships between gene function and fitness. Experiments to test targets and agents of natural selection within a genomic context are necessary for identifying the adaptive consequences of individual alleles.