Identification of peptide ligands to the chemokine receptor CCR5 and their maturation by gene shuffling.

The determination of protein-protein interactions and their role in diverse pathophysiological processes is a promising approach to the identification of molecules of therapeutic potential. This paper describes the identification of peptidic CCR5 receptor ligands as potential drug leads against HIV-1 infection using in vitro evolution based on phage display. A phage-displayed peptide library was used to select for anti-CCR5 peptide. Further in vitro evolution of the peptide by exon shuffling was performed to identify peptides with optimized characteristics for CCR5 receptor. This peptide inhibited HIV coreceptor activity in a cell fusion assay with an IC50 of 5 microM. It did not exhibit either agonistic or antagonistic activity on CCR5 in the concentration range used. To our knowledge, this is a first report that describes the identification of peptide ligands specific to the CCR5 receptor from a phage-displayed library and the maturation of the selected peptide sequence by gene shuffling.

Using parthenogenetic lineages to identify advantages of sex

The overwhelming predominance of sexual reproduction in nature is surprising given that sex is expected to confer profound costs in terms of production of males and the breakup of beneficial allele combinations. Recognition of these theoretical costs was the inspiration for a large body of empirical research-typically focused on comparing sexual and asexual organisms, lineages, or genomes-dedicated to identifying the advantages and maintenance of sex in natural populations. Despite these efforts, why sex is so common remains unclear. Here, we argue that we can generate general insights into the advantages of sex by taking advantage of parthenogenetic taxa that differ in such characteristics as meiotic versus mitotic offspring production, ploidy level, and single versus multiple and hybrid versus non-hybrid origin. We begin by evaluating benefits that sex can confer via its effects on genetic linkage, diversity, and heterozygosity and outline how the three classes of benefits make different predictions for which type of parthenogenetic lineage would be favored over others. Next, we describe the type of parthenogenetic model system (if any) suitable for testing whether the hypothesized benefit might contribute to the maintenance of sex in natural populations, and suggest groups of organisms that fit the specifications. We conclude by discussing how empirical estimates of characteristics such as time since derivation and number of independent origins of asexual lineages from sexual ancestors, ploidy levels, and patterns of molecular evolution from representatives of these groups can be used to better understand which mechanisms maintain sex in natural populations.

Vitellogenin Underwent Subfunctionalization to Acquire Caste and Behavioral Specific Expression in the Harvester Ant Pogonomyrmex barbatus.

The reproductive ground plan hypothesis (RGPH) proposes that the physiological pathways regulating reproduction were co-opted to regulate worker division of labor. Support for this hypothesis in honeybees is provided by studies demonstrating that the reproductive potential of workers, assessed by the levels of vitellogenin (Vg), is linked to task performance. Interestingly, contrary to honeybees that have a single Vg ortholog and potentially fertile nurses, the genome of the harvester ant Pogonomyrmex barbatus harbors two Vg genes (Pb_Vg1 and Pb_Vg2) and nurses produce infertile trophic eggs. P. barbatus, thus, provides a unique model to investigate whether Vg duplication in ants was followed by subfunctionalization to acquire reproductive and non-reproductive functions and whether Vg reproductive function was co-opted to regulate behavior in sterile workers. To investigate these questions, we compared the expression patterns of P. barbatus Vg genes and analyzed the phylogenetic relationships and molecular evolution of Vg genes in ants. qRT-PCRs revealed that Pb_Vg1 is more highly expressed in queens compared to workers and in nurses compared to foragers. By contrast, the level of expression of Pb_Vg2 was higher in foragers than in nurses and queens. Phylogenetic analyses show that a first duplication of the ancestral Vg gene occurred after the divergence between the poneroid and formicoid clades and subsequent duplications occurred in the lineages leading to Solenopsis invicta, Linepithema humile and Acromyrmex echinatior. The initial duplication resulted in two Vg gene subfamilies preferentially expressed in queens and nurses (subfamily A) or in foraging workers (subfamily B). Finally, molecular evolution analyses show that the subfamily A experienced positive selection, while the subfamily B showed overall relaxation of purifying selection. Our results suggest that in P. barbatus the Vg gene underwent subfunctionalization after duplication to acquire caste- and behavior- specific expression associated with reproductive and non-reproductive functions, supporting the validity of the RGPH in ants.

Can we continue to neglect genomic variation in introgression rates when inferring the history of speciation? A case study in a Mytilus hybrid zone.

The use of molecular data to reconstruct the history of divergence and gene flow between populations of closely related taxa represents a challenging problem. It has been proposed that the long-standing debate about the geography of speciation can be resolved by comparing the likelihoods of a model of isolation with migration and a model of secondary contact. However, data are commonly only fit to a model of isolation with migration and rarely tested against the secondary contact alternative. Furthermore, most demographic inference methods have neglected variation in introgression rates and assume that the gene flow parameter (Nm) is similar among loci. Here, we show that neglecting this source of variation can give misleading results. We analysed DNA sequences sampled from populations of the marine mussels, Mytilus edulis and M. galloprovincialis, across a well-studied mosaic hybrid zone in Europe and evaluated various scenarios of speciation, with or without variation in introgression rates, using an Approximate Bayesian Computation (ABC) approach. Models with heterogeneous gene flow across loci always outperformed models assuming equal migration rates irrespective of the history of gene flow being considered. By incorporating this heterogeneity, the best-supported scenario was a long period of allopatric isolation during the first three-quarters of the time since divergence followed by secondary contact and introgression during the last quarter. By contrast, constraining migration to be homogeneous failed to discriminate among any of the different models of gene flow tested. Our simulations thus provide statistical support for the secondary contact scenario in the European Mytilus hybrid zone that the standard coalescent approach failed to confirm. Our results demonstrate that genomic variation in introgression rates can have profound impacts on the biological conclusions drawn from inference methods and needs to be incorporated in future studies.

SlimCodeML: An Optimized Version of CodeML for the Branch-Site Model

CodeML (part of the PAML package) im- plements a maximum likelihood-based approach to de- tect positive selection on a specific branch of a given phylogenetic tree. While CodeML is widely used, it is very compute-intensive. We present SlimCodeML, an optimized version of CodeML for the branch-site model. Our performance analysis shows that SlimCodeML substantially outperforms CodeML (up to 9.38 times faster), especially for large-scale genomic analyses.

Rolf Bernander (1956-2014): pioneer of the archaeal cell cycle.

On 19 January 2014 Rolf ('Roffe') Bernander passed away unexpectedly. Rolf was a dedicated scientist; his research aimed at unravelling the cell biology of the archaeal domain of life, especially cell cycle-related questions, but he also made important contributions in other areas of microbiology. Rolf had a professor position in the Molecular Evolution programme at Uppsala University, Sweden for about 8 years, and in January 2013 he became chair professor at the Department of Molecular Biosciences, The Wenner-Gren Institute at Stockholm University in Sweden. Rolf was an exceptional colleague and will be deeply missed by his family and friends, and the colleagues and co-workers that he leaves behind in the scientific community. He will be remembered for his endless enthusiasm for science, his analytical mind, and his quirky sense of humour.

The role of climatic tolerances and seed traits in reduced extinction rates of temperate polygonaceae.

The latitudinal diversity gradient (LDG) is one of the most striking and consistent biodiversity patterns across taxonomic groups. We investigate the species richness gradient in the buckwheat family, Polygonaceae, which exhibits a reverse LDG and is, thus, decoupled from dominant gradients of energy and environmental stability that increase toward the tropics and confound mechanistic interpretations. We test competing age and evolutionary diversification hypotheses, which may explain the diversification of this plant family over the past 70 million years. Our analyses show that the age hypothesis, which posits that clade richness is positively correlated with the ecological and evolutionary time since clade origin, fails to explain the richness gradient observed in Polygonaceae. However, an evolutionary diversification hypothesis is highly supported, with diversification rates being 3.5 times higher in temperate clades compared to tropical clades. We demonstrate that differences in rates of speciation, migration, and molecular evolution insufficiently explain the observed patterns of differential diversification rates. We suggest that reduced extinction rates in temperate clades may be associated with adaptive responses to selection, through which seed morphology and climatic tolerances potentially act to minimize risk in temporally variable environments. Further study is needed to understand causal pathways among these traits and factors correlated with latitude.

Patterns of positive selection in seven ant genomes.

The evolution of ants is marked by remarkable adaptations that allowed the development of very complex social systems. To identify how ant-specific adaptations are associated with patterns of molecular evolution, we searched for signs of positive selection on amino-acid changes in proteins. We identified 24 functional categories of genes which were enriched for positively selected genes in the ant lineage. We also reanalyzed genome-wide data sets in bees and flies with the same methodology to check whether positive selection was specific to ants or also present in other insects. Notably, genes implicated in immunity were enriched for positively selected genes in the three lineages, ruling out the hypothesis that the evolution of hygienic behaviors in social insects caused a major relaxation of selective pressure on immune genes. Our scan also indicated that genes implicated in neurogenesis and olfaction started to undergo increased positive selection before the evolution of sociality in Hymenoptera. Finally, the comparison between these three lineages allowed us to pinpoint molecular evolution patterns that were specific to the ant lineage. In particular, there was ant-specific recurrent positive selection on genes with mitochondrial functions, suggesting that mitochondrial activity was improved during the evolution of this lineage. This might have been an important step toward the evolution of extreme lifespan that is a hallmark of ants.

Phylogenomics of C(4) photosynthesis in sedges (Cyperaceae): multiple appearances and genetic convergence.

C(4) photosynthesis is an adaptive trait conferring an advantage in warm and open habitats. It originated multiple times and is currently reported in 18 plant families. It has been recently shown that phosphoenolpyruvate carboxylase (PEPC), a key enzyme of the C(4) pathway, evolved through numerous independent but convergent genetic changes in grasses (Poaceae). To compare the genetics of multiple C(4) origins on a broader scale, we reconstructed the evolutionary history of the C(4) pathway in sedges (Cyperaceae), the second most species-rich C(4) family. A sedge phylogeny based on two plastome genes (rbcL and ndhF) has previously identified six fully C(4) clades. Here, a relaxed molecular clock was used to calibrate this tree and showed that the first C(4) acquisition occurred in this family between 19.6 and 10.1 Ma. According to analyses of PEPC-encoding genes (ppc), at least five distinct C(4) origins are present in sedges. Two C(4) Eleocharis species, which were unrelated in the plastid phylogeny, acquired their C(4)-specific PEPC genes from a single source, probably through reticulate evolution or a horizontal transfer event. Acquisitions of C(4) PEPC in sedges have been driven by positive selection on at least 16 codons (3.5% of the studied gene segment). These sites underwent parallel genetic changes across the five sedge C(4) origins. Five of these sites underwent identical changes also in grass and eudicot C(4) lineages, indicating that genetic convergence is most important within families but that identical genetic changes occurred even among distantly related taxa. These lines of evidence give new insights into the constraints that govern molecular evolution.

Genetic regulation of colony social organization in fire ants: an integrative overview

Expression of colony social organization in fire ants appears to be under the control of a single Mendelian factor of large effect. Variation in colony queen number in Solenopsis invicta and its relatives is associated with allelic variation at the gene Gp-9, but not with variation at other unlinked genes; workers regulate queen identity and number on the basis of Gp-9 genotypic compatibility. Nongeneticfactors, such as prior social experience, queen reproductive status, and local environment, have negligible effects on queen number which illustrates the nearly complete penetrance of Gp-9. As predicted, queen number can be manipulated experimentally by altering worker Gp-9 genotype frequencies. The Gp-9 allele lineage associated with polygyny in South American fire? ants has been retained across multiple speciation events, which may signal the action of balancing selection to maintain social polymorphism in these species. Moreover positive selection is implicated in driving the molecular evolution of Gp-9 in association with the origin of polygyny. The identity of the product of Gp-9 as an odorant-binding protein suggests plausible scenarios for its direct involvement in the regulation of queen number via a role in chemical communication. While these and other lines of evidence show that Gp-9 represents a legitimate candidate gene of major effect, studies aimed at determining (i) the biochemical pathways in which GP-9 functions; (ii) the phenotypic effects of molecular variation at Gp-9 and other pathway genes; and (iii) the potential involvement of genes in linkage disequilibrium with Gp-9 are needed to elucidate the genetic architecture underlying social organization in fire ants. Information that reveals the links between molecular variation, individual phenotype, and colony-level behaviors, combined with behavioral models that incorporate details of the chemical communication involved in regulating queen number will yield a novel integrated view of the evolutionary changes underlying a key social adaptation.

The distribution of the dinucleotide CpG and cytosine methylation in the vitellogenin gene family.

Sequence data from regions of five vertebrate vitellogenin genes were used to examine the frequency, distribution, and mutability of the dinucleotide CpG, the preferred modification site for eukaryotic DNA methyltransferases. The observed level of the CpG dinucleotide in all five genes was markedly lower than that expected from the known mononucleotide frequencies. CpG suppression was greater in introns than in exons. CpG-containing codons were found to be avoided in the vitellogenin genes, but not completely despite the redundancy of the genetic code. Frequency and distribution patterns of this dinucleotide varied dramatically among these otherwise closely related genes. Dense clusters of CpG dinucleotides tended to appear in regions of either functional or structural interest (e.g., in the transposon-like Vi-element of Xenopus) and these clusters contained 5-methylcytosine (5 mC). 5 mC is known to undergo deamination to form thymidine, but the extent to which this transition occurs in the heavily methylated genomes of vertebrates and its contribution to CpG suppression are still unclear. Sequence comparison of the methylated vitellogenin gene regions identified C----T and G----A substitutions that were found to occur at relatively high frequencies. The predicted products of CpG deamination, TpG and CpA, were elevated. These findings are consistent with the view that CpG distribution and methylation are interdependent and that deamination of 5 mC plays an important role in promoting evolutionary change at the nucleotide sequence level.

The hourglass and the early conservation models--co-existing patterns of developmental constraints in vertebrates.

Developmental constraints have been postulated to limit the space of feasible phenotypes and thus shape animal evolution. These constraints have been suggested to be the strongest during either early or mid-embryogenesis, which corresponds to the early conservation model or the hourglass model, respectively. Conflicting results have been reported, but in recent studies of animal transcriptomes the hourglass model has been favored. Studies usually report descriptive statistics calculated for all genes over all developmental time points. This introduces dependencies between the sets of compared genes and may lead to biased results. Here we overcome this problem using an alternative modular analysis. We used the Iterative Signature Algorithm to identify distinct modules of genes co-expressed specifically in consecutive stages of zebrafish development. We then performed a detailed comparison of several gene properties between modules, allowing for a less biased and more powerful analysis. Notably, our analysis corroborated the hourglass pattern at the regulatory level, with sequences of regulatory regions being most conserved for genes expressed in mid-development but not at the level of gene sequence, age, or expression, in contrast to some previous studies. The early conservation model was supported with gene duplication and birth that were the most rare for genes expressed in early development. Finally, for all gene properties, we observed the least conservation for genes expressed in late development or adult, consistent with both models. Overall, with the modular approach, we showed that different levels of molecular evolution follow different patterns of developmental constraints. Thus both models are valid, but with respect to different genomic features.

K-r/K-c but not d(N)/d(S) correlates positively with body mass in birds, raising implications for inferring lineage-specific selection

Background: The ratio of the rates of non-synonymous and synonymous substitution (d(N)/d(S)) is commonly used to estimate selection in coding sequences. It is often suggested that, all else being equal, d(N)/d(S) should be lower in populations with large effective size (Ne) due to increased efficacy of purifying selection. As N-e is difficult to measure directly, life history traits such as body mass, which is typically negatively associated with population size, have commonly been used as proxies in empirical tests of this hypothesis. However, evidence of whether the expected positive correlation between body mass and d(N)/d(S) is consistently observed is conflicting. Results: Employing whole genome sequence data from 48 avian species, we assess the relationship between rates of molecular evolution and life history in birds. We find a negative correlation between dN/dS and body mass, contrary to nearly neutral expectation. This raises the question whether the correlation might be a method artefact. We therefore in turn consider non-stationary base composition, divergence time and saturation as possible explanations, but find no clear patterns. However, in striking contrast to d(N)/d(S), the ratio of radical to conservative amino acid substitutions (K-r/K-c) correlates positively with body mass. Conclusions: Our results in principle accord with the notion that non-synonymous substitutions causing radical amino acid changes are more efficiently removed by selection in large populations, consistent with nearly neutral theory. These findings have implications for the use of d(N)/d(S) and suggest that caution is warranted when drawing conclusions about lineage-specific modes of protein evolution using this metric.

Amphibian albumins as members of the albumin, alpha-fetoprotein, vitamin D-binding protein multigene family.

The Xenopus laevis 68-kd and 74-kd albumin amino acid sequences are examined with respect to their relationship to the other known members of the albumin/alpha-fetoprotein/vitamin D-binding protein gene family. Each of the three members of this family presents a unique pattern of conserved regions indicating a differential selective pressure related to specific functional characteristics. Furthermore, an evolutionary tree of these genes was deduced from the divergence times calculated from direct nucleotide sequence comparisons of individual gene pairs. These calculations indicate that the vitamin D-binding protein/albumin separation occurred 560-600 million years (Myr) ago and the albumin/alpha-fetoprotein divergence 280 Myr ago. This observation leads to the hypothesis according to which the albumin/alpha-fetoprotein gene duplication occurred shortly after the amphibian/reptile separation. Consequently, and unlike mammals, amphibians and fishes should lack an alpha-fetoprotein in their serum at larval stages, which is consistent with a recent analysis of serum proteins in Xenopus laevis larvae. This hypothesis now will have to be tested further in additional lower vertebrates.

Species-specific population structure in rock-specialized sympatric cichlid species in Lake Tanganyika, East Africa.

Species richness and geographical phenotypic variation in East African lacustrine cichlids are often correlated with ecological specializations and limited dispersal. This study compares mitochondrial and microsatellite genetic diversity and structure among three sympatric rock-dwelling cichlids of Lake Tanganyika, Eretmodus cyanostictus, Tropheus moorii, and Ophthalmotilapia ventralis. The species represent three endemic, phylogenetically distinct tribes (Eretmodini, Tropheini, and Ectodini), and display divergent ecomorphological and behavioral specialization. Sample locations span both continuous, rocky shoreline and a potential dispersal barrier in the form of a muddy bay. High genetic diversity and population differentiation were detected in T. moorii and E. cyanostictus, whereas much lower variation and structure were found in O. ventralis. In particular, while a 7-km-wide muddy bay curtails dispersal in all three species to a similar extent, gene flow along mostly continuous habitat appeared to be controlled by distance in E. cyanostictus, further restricted by site philopatry and/or minor habitat discontinuities in T. moorii, and unrestrained in O. ventralis. In contrast to the general pattern of high gene flow along continuous shorelines in rock-dwelling cichlids of Lake Malawi, our study identifies differences in population structure among stenotopic Lake Tanganyika species. The amount of genetic differentiation among populations was not related to the degree of geographical variation of body color, especially since more phenotypic variation is observed in O. ventralis than in the genetically highly structured E. cyanostictus.