Molecular Biology and Evolution - Advance Access

Phylogenetic analysis of gene structure and alternative splicing in {alpha}-actinins

Lek, M., MacArthur, D. G., Yang, N., North, K. N. — Fri, 06 Nov 2009 02:40:29 PST

The -actinins are an important family of actin-binding proteins with the ability to cross-link actin filaments when in dimer form. Members of the -actinin family share a domain topology composed of highly conserved actin binding and EF-hand domains separated by a rod domain composed of spectrin-like repeats. Functional diversity within this family has arisen through exon duplication and the formation of alternate splice isoforms as well as gene duplications during the evolution of vertebrates. In addition to the known functional domains, -actinins also contain a consensus PDZ binding site. The completed genome sequence of over 32 invertebrate species has allowed the analysis of gene structure and exon/gene duplication over a diverse range of phyla. Our analysis shows that relative to early branching metazoans, considerable intron loss especially in arthropods with few cases of intron gains. The C-terminal PDZ binding site is conserved in nearly all invertebrates but is missing in some nematodes and platyhelminths. Alternative splicing in the actin-binding domain is conserved in chordates, arthropods and some nematodes and platyhelminths. In contrast, alternative splicing of the EF-hand domain is only observed in chordates. Finally, given the prevalence of exon duplications seen in the actin-binding domain, this may act as a significant mechanism in the modification of actin binding properties.

Intragenic Recombination Events and Evidence for Hybrid Speciation in Nicotiana (Solanaceae)

Fri, 06 Nov 2009 02:40:28 PST

Reticulate evolution may function both at the species-level, through homoploid and polyploid hybridization, and below the species level, through inter- and intragenic recombination. These processes represent challenges for the reconstruction of evolutionary relationships between species, because they cannot be represented adequately with bifurcating trees. We use data from low-copy nuclear genes to evaluate long-standing hypotheses of homoploid (interspecific) hybrid speciation in Nicotiana (Solanaceae) and reconstruct a complex series of reticulation events that have been important in the evolutionary history of this genus. Hybrid origins for three diploid species (N. glauca, N. linearis and N. spegazzinii) are inferred on the basis of gene tree incongruence, evidence for inter-allelic recombination between likely parental alleles, and support for incompatible splits in Lento plots. Phylogenetic analysis of recombinant gene sequences illustrates that recombinants may be resolved with one of their progenitor lineages with a high posterior probability under Bayesian inference, and thus there is no indication of the conflict between phylogenetic signals that results from reticulation. Our results illustrate the importance of hybridization in shaping evolution in Nicotiana and also show that intragenic recombination may be relatively common. This finding demonstrates that it is important to investigate the possibility of recombination when aiming to detect hybrids from DNA sequence data and reconstruct patterns of reticulate evolution between species.

Large-scale parsimony analysis of metazoan indels in protein-coding genes

Belinky, F., Cohen, O., Huchon, D. — Wed, 28 Oct 2009 08:29:22 PDT

Insertions and deletions (indels) are considered to be rare evolutionary events, the analysis of which may resolve controversial phylogenetic relationships. Indeed, indel characters are often assumed to be less homoplastic than amino-acid and nucleotide substitutions and, consequently, more reliable markers for phylogenetic reconstruction. In this study we analyzed indels from over a thousand metazoan orthologous genes. We studied the impact of different species sampling, ortholog datasets, lengths of included indels, and indel-coding methods, on the resulting metazoan tree. Our results show that, similar to sequence substitutions, indels are homoplastic characters and their analysis is sensitive to the long-branch attraction artifact. Furthermore, improving the taxon sampling and choosing a closely-related outgroup greatly impact the phylogenetic inference. Our indel-based inferences support the Ecdysozoa hypothesis over the Coelomata hypothesis, and suggest that sponges are a sister clade to other animals.

Evolutionary Fingerprinting of Genes

Wed, 28 Oct 2009 08:29:20 PDT

Over time, natural selection molds every gene into a unique mosaic of sites evolving rapidly or resisting change - an ‘evolutionary fingerprint’ of the gene. Aspects of this evolutionary fingerprint, such as the site-specific ratio of nonsynonymous to synonymous substitution rates (dN/dS), are commonly used to identify genetic features of potential biological interest; however, no framework exists for comparing evolutionary fingerprints between genes. We hypothesize that protein coding genes with similar protein structure and/or function tend to have similar evolutionary fingerprints, and that comparing evolutionary fingerprints can be useful for discovering similarities between genes in a way that is analogous to, but independent of, discovery of similarity via sequence-based comparison tools such as BLAST.

To test this hypothesis, we develop a novel model of coding sequence evolution that uses a general bivariate discrete parameterization of the evolutionary rates. We show that this approach provides a better fit to the data using a smaller number of parameters than existing models. Next, we use the model to represent evolutionary fingerprints as probability distributions and present a methodology for comparing these distributions in a way that is robust against variations in data set size and divergence. Finally, using sequences of three rapidly evolving RNA viruses (HIV-1, Hepatitis C virus and Influenza A virus) we demonstrate that genes within the same functional group tend to have similar evolutionary fingerprints. Our framework provides a sound statistical foundation for efficient inference and comparison of evolutionary rate patterns in arbitrary collections of gene alignments, clustering homologous and non-homologous genes and investigation of biological and functional correlates of evolutionary rates.

Phylodynamics of HIV-1 from a Phase III AIDS vaccine trial in North America

Wed, 28 Oct 2009 08:29:19 PDT

In 2003, a phase III placebo-controlled trial (VAX004) of a candidate HIV-1 vaccine (AIDSVAX B/B) was completed in 5,403 volunteers at high risk for HIV-1 infection from North America and the Netherlands. A total of 368 individuals became infected with HIV-1 during the trial. The envelope glycoprotein gene (gp120) from the HIV-1 subtype B viruses infecting 349 patients was sequenced from clinical samples taken as close as possible to the time of diagnosis, rendering a final data set of 1,047 sequences (1,032 from North America and 15 from the Netherlands). Here, we used these data in combination with other sequences available in public databases to assess HIV-1 variation as a function of vaccination treatment, geographic region, race, risk behavior, and viral load. Viral samples did not show any phylogenetic structure for any of these factors, but individuals with different viral loads showed significant differences (P = 0.009) in genetic diversity. The estimated time of emergence of HIV-1 subtype B in USA was around 1966-1970. Despite the fact that the number of AIDS cases has decreased in North America since the early nineties, HIV-1 genetic diversity seems to have remained almost constant over time. This study represents one of the largest molecular epidemiologic surveys of viruses responsible for new HIV-1 infections in North America and could help the selection of epidemiologically representative vaccine antigens to include in the next generation of candidate HIV-1 vaccines.

An explicit signature of balancing selection for color vision variation in New World monkeys

Tue, 27 Oct 2009 12:16:26 PDT

Color vision is an important characteristic of primates and, intriguingly, Neotropical monkeys are highly polymorphic for this trait. Recent field studies have challenged the conventional view that trichromatic color vision is more adaptive than dichromatic color vision. No study has investigated the pattern of genetic variation in the long to middle-wavelength sensitive (L-M or red-green) opsin gene as compared to that of other genomic regions (neutral references) in wild populations of New World monkeys to look for the signature of natural selection. Here we report such a study conducted on spider monkeys and capuchin monkeys inhabiting Santa Rosa National Park, Costa Rica. The nucleotide sequence of the L-M opsin gene was more polymorphic than the sequences of the neutral references, although the opsin gene sequences were not more divergent between the two species than were the sequences of the neutral references. In a coalescence simulation that took into account the observed nucleotide diversity of the neutral references, the Tajima's D value of the L-M opsin gene deviated significantly in a positive direction from the expected range. These results are the first to statistically demonstrate balancing selection acting on the polymorphic L-M opsin gene of New World monkeys. Taking the results of behavioral and genetic studies together, the balancing selection we detected may indicate that coexistence of different color vision types in the same population, also characteristic of humans, is adaptive.

Bacterial genetic signatures of human social phenomena among M.tuberculosis from an Aboriginal Canadian population

Pepperell, C., Hoeppner, V. H., Lipatov, M., Schoolnik, G. K., Feldman, M. W. — Tue, 27 Oct 2009 12:16:24 PDT

Despite a widespread global distribution and highly variable disease phenotype, there is little DNA sequence diversity among isolates of Mycobacterium tuberculosis. In addition, many regional population genetic surveys have revealed a stereotypical structure in which a single clone, lineage or clade makes up the majority of the population. It is often assumed that dominant clones are highly adapted; that is, the overall structure of M.tb populations is the result of positive selection. In order to test this assumption, we analyzed genetic data from extant populations of bacteria circulating in Aboriginal communities in Saskatchewan, Canada. Demographic parameters of the bacterial population were estimated from archival epidemiological data collected over ~130 years since the onset of epidemic tuberculosis in the host communities. Bacterial genetic data were tested against neutral theory expectations and the local evolutionary history of M.tb investigated by phylogenetic analysis. Our findings are not consistent with positive selection on the bacterial population. Instead, we uncovered founder effects persisting over decades, and barriers to gene flow within the bacterial population. Simulation experiments suggested that a combination of these neutral influences could result in the stereotypical structure of M.tb populations. Some aspects of population structure were suggestive of background selection and data were on the whole consistent with combined effects of population bottlenecks, subdivision and background selection. Neutral phenomena, namely bottlenecks and partitions within populations, are prominent influences on the evolution of M.tuberculosis and likely contribute to restricted genetic diversity observed within this species. Given these influences, a complex evolutionary model will be required to define the relative fitness of different M.tb lineages and, ultimately, to uncover the genetic basis for its success as a pathogen.

Vertebrate Rhodopsin Adaptation to Dim Light via Rapid Meta-II Intermediate Formation

Sugawara, T., Imai, H., Nikaido, M., Imamoto, Y., Okada, N. — Sun, 25 Oct 2009 19:33:30 PDT

Rhodopsin is a photoreceptive protein present in vertebrate rod photoreceptor cells, which are responsible for scotopic vision. Recent molecular studies have shown that several aquatic vertebrate species have independently acquired rhodopsin containing Asp83Asn, Glu122Gln and Ala292Ser substitutions, causing a blue-shift in the rhodopsin absorption spectra for adaptation to the blue-green photic environment in deep water. Here, we provide new evidence for the evolutionary and functional relevance of the Asp83Asn substitution. Spectroscopic and kinetic analyses of rhodopsins in six cichlid fishes from the East African Great Lakes using charge-coupled device spectrophotometer revealed that the Asp83Asn substitution accelerated formation of meta-II, a rhodopsin intermediate crucial for activation of the G-protein transducin. Because rapid formation of meta-II likely results in effective transduction of photic signals, it is reasonable to assume that deep-water cichlid species have acquired rhodopsin containing Asn83 to adapt to dim lighting. Remarkably, rhodopsin containing Asn83 has been identified in terrestrial vertebrates such as bats, and these rhodopsin variants also exhibit accelerated meta-II formation. Our results indicated that the Asp83Asn substitution observed in a variety of animal species was acquired independently in many different lineages during vertebrate evolution for adaptation to dimly lit environments.

SeaView version 4 : a multiplatform graphical user interface for sequence alignment and phylogenetic tree building

Gouy, M., Guindon, S., Gascuel, O. — Fri, 23 Oct 2009 02:56:10 PDT

We present SeaView version 4, a multiplatform program designed to facilitate multiple alignment and phylogenetic tree building from molecular sequence data through the use of a graphical user interface. SeaView version 4 combines all the functions of the widely used programs SeaView (in its previous versions) and Phylo_win, and expands them by adding network access to sequence databases, alignment with arbitrary algorithm, maximum likelihood tree-building with PhyML, and display, printing, and copy-to-clipboard of rooted or unrooted, binary or multifurcating phylogenetic trees. In relation to the wide present offer of tools and algorithms for phylogenetic analyses, SeaView is especially useful for teaching and for occasional users of such software. SeaView is freely available at http://pbil.univ-lyon1.fr/software/seaview.

Evidence that localised variation in primate sequence divergence arises from an influence of nucleosome placement on DNA repair

Ying, H., Epps, J., Williams, R., Huttley, G. — Tue, 20 Oct 2009 07:48:36 PDT

Understanding the origins of localised substitution rate heterogeneity has important implications for identifying functional genomic sequences. Outside of gene regions, the origins of rate heterogeneity remain unclear. Experimental studies establish that chromatin compaction affects rates of both DNA lesion formation and repair. A functional association between chromatin status and 5-methyl-cytosine also exists. These suggest that both the total rate and type of substitution will be affected by chromatin status. Regular positioning of nucleosomes, the building block of chromatin, further predicts that substitution rate and type should vary spatially in an oscillating manner. We addressed chromatin's influence on substitution rate and type in primates. Matched numbers of sites were sampled from Dnase I Hypersensitive (DHS) and closed chromatin control flank (Flank). Likelihood ratio tests revealed significant excesses of total and of transition substitutions in Flank compared with matched DHS for both intergenic and intronic samples. An additional excess of CpG transitions was evident for the intergenic, but not intronic, regions. Fluctuation in substitution rate along ~ 1800 primate promoters were measured using phylogenetic footprinting. Significant positive correlations were evident between the substitution rate and a nucleosome score from resting human T-cells, with up to ~ 50% of the variance in substitution rate accounted for. Using signal processing techniques, a dominant oscillation at ~ 200 bp was evident in both the substitution rate and nucleosome score. Our results support a role for differential DNA repair rates between open and closed chromatin in the spatial distribution of rate heterogeneity.

Natural selection on synonymous and non-synonymous mutations shape patterns of polymorphism in Populus tremula

Ingvarsson, P. K. — Fri, 16 Oct 2009 07:30:31 PDT

One important goal of population genetics is to understand the relative importance of different evolutionary processes for shaping variation in natural populations. Here I use multilocus data to show that natural selection on both synonymous and non-synonymous mutations plays an important role in shaping levels of synonymous polymorphism in European aspen (Populus tremula). Previous studies have documented a preferential fixation of synonymous mutations encoding preferred codons in P. tremula. The results presented here show that this has resulted in an increase in codon bias in P. tremula, consistent with stronger selection acting on synonymous codon usage. In addition, positive selection on non-synonymous mutations appear to be common in P. tremula, with approximately 30% of all mutations having been fixed by positive selection. In addition, the recurrent fixation of beneficial mutations also reduce standing levels of polymorphism as evidenced by a significantly negative relationship between the rate of protein evolution synonymous site diversity and silent site diversity. Finally, I use Approximate Bayesian methods to estimate the strength of selection acting on beneficial substitutions. These calculations show that recurrent hitchhiking reduce polymorphism by, on average, 30%. The product of strength of selection acting of beneficial mutations and the rate by which these occur across the genome (2N_es) equals 1.54 x 10^–7, which is in line with estimates from Drosophila where recurrent hitchhiking also have been shown to have significant effects on standing levels of polymorphism.

Novel venom proteins produced by differential domain-expression strategies in Beaded lizards and Gila monsters (genus Heloderma)

Thu, 15 Oct 2009 19:22:37 PDT

The origin and evolution of venom proteins in helodermatid lizards was investigated by multidisciplinary techniques. Our analyses elucidated novel toxin types resultant from three unique domain-expression processes: i) the first full-length sequences of Lethal Toxin isoforms (helofensins) revealed this toxin type to be constructed by an ancestral mono-domain, mono-product gene (beta-defensin) which underwent three tandem domain duplications to encode a tetra-domain, mono-product with a possible novel protein fold; (ii) an ancestral mono-domain gene (encoding a natriuretic peptide) was medially extended to become a penta-domain, penta-product through the additional encoding of four tandemly repeated proline-rich peptides (helokinestatins), with the five discrete peptides liberated from each other by post-translational proteolysis; and iii) an ancestral multi-domain, multi-product gene belonging to the VIP/glucagon family being mutated to encode for a mono-domain, mono-product (exendins) followed by duplication and diversification into two variant classes (exendins 1&2 and exendins 3&4). Bioactivity characterization of exendin and helokinestatin elucidated variable cardioactivity between isofroms within each class. These results highlight the importance of utilising evolutionary-based search strategies for biodiscovery and the virtually unexplored potential of lizard venoms in drug design and discovery.

Fast and consistent estimation of species trees using supermatrix rooted triples

DeGiorgio, M., Degnan, J. H. — Thu, 15 Oct 2009 06:32:04 PDT

Concatenated sequence alignments are often used to infer species-level relationships. Previous studies have shown that analysis of concatenated data using maximum likelihood (ML) can produce misleading results when loci have differing gene tree topologies due to incomplete lineage sorting. Here, we develop a polynomial-time method that utilizes the modified mincut supertree algorithm to construct an estimated species tree from inferred rooted triples of concatenated alignments. We term this method SuperMatrix Rooted Triple (SMRT) and use the notation SMRT-ML when rooted triples are inferred by ML. We use simulations to investigate the performance of SMRT-ML under Jukes-Cantor and General Time-Reversible substitution models for four- and five-taxon species trees, and also apply the method to an empirical dataset of yeast genes. We find that SMRT-ML converges to the correct species tree in many cases in which maximum likelihood on the full concatenated dataset fails to do so. SMRT-ML can be conservative in that its output tree is often partially unresolved for problematic clades. We show analytically that when the species tree is clocklike and mutations occur under the Cavender-Farris-Neyman substitution model, as the number of genes increases, SMRT-ML is increasingly likely to infer the correct species tree even when the most likely gene tree does not match the species tree. SMRT-ML is therefore a computationally efficient and statistically consistent estimator of the species tree when gene trees are distributed according to the multispecies coalescent model.

Indel-associated mutation rate varies with mating system in flowering plants

Hollister, J. D., Ross-Ibarra, J., Gaut, B. S. — Tue, 13 Oct 2009 05:37:29 PDT

A recently proposed mutational mechanism, indel-associated mutation (IDAM), posits that heterozygous insertions/deletions (indels) increase the point mutation rate at nearby nucleotides due to errors during meiosis. This mechanism could have especially dynamic consequences for the evolution of plant genomes, because the high degree of variation in the rate of self-fertilization among plant species causes differences in the heterozygosity of alleles, including indel alleles, segregating in plant species. In this study, we investigated the consequences of IDAM for species differing in mating system using both forward population genetic simulations and genome-wide DNA re-sequencing data from Arabidopsis thaliana, Oryza sativa, and Oryza rufipogon. Simulations of different levels of selfing suggest that the effect of IDAM on surrounding nucleotide diversity should decrease with increasing selfing rate. Further simulations incorporating selfing rates and the time of onset of selfing suggest that the time since the switch to selfing also affects patterns of nucleotide diversity due to IDAM. Population genetic analyses of A. thaliana and Oryza DNA sequence datasets empirically confirmed our simulation results, revealing the strongest effect of IDAM in the out-crossing O. rufipogon, a weaker effect in the recently evolved selfer O. sativa, and the weakest effect in the relatively ancient selfer A. thaliana. These results support the novel idea that differences in life-history, such as the level of selfing, can affect the per-individual mutation rate among species.

A Dirichlet process covarion mixture model and its assessments using posterior predictive discrepancy tests

Zhou, Y., Brinkmann, H., Rodrigue, N., Lartillot, N., Philippe, H. — Mon, 12 Oct 2009 08:03:15 PDT

Heterotachy, the variation of substitution rate at a site across time, is a prevalent phenomenon in nucleotide and amino acid alignments, which may mislead probabilistic-based phylogenetic inferences. The covarion model is a special case of heterotachy, in which sites change between the "ON" state (allowing substitutions according to any particular model of sequence evolution) and the "OFF" state (prohibiting substitutions). In current implementations, the switch rates between ON and OFF states are homogeneous across sites, a hypothesis that has never been tested. In this study we developed an infinite mixture model, called the covarion mixture (CM) model, which allows the covarion parameters to vary across sites, controlled by a Dirichlet process prior. Moreover, we combine the covarion mixture model with other approaches. We use a second independent Dirichlet process that models the heterogeneities of amino acid equilibrium frequencies across sites, known as the CAT model, and general rate-across-site heterogeneity is modeled by a gamma distribution. The application of the CM model to several large alignments demonstrates that the covarion parameters are significantly heterogeneous across sites. We describe posterior predictive discrepancy tests, and use these to demonstrate the importance of these different elements of the models.

A worldwide survey of human male demographic history based on Y-SNP and Y-STR data from the HGDP-CEPH populations

Mon, 12 Oct 2009 08:03:12 PDT

We have investigated human male demographic history using 590 males from 51 populations in the HGDP-CEPH worldwide panel, typed with 37 Y-SNPs and 65 Y-STRs, and analyzed with the program BATWING. The general patterns we observe show a gradient from the oldest population TMRCAs and expansion times together with the largest effective population sizes in Africa, to the youngest times and smallest effective population sizes in the Americas. These parameters are significantly negatively correlated with distance from East Africa and the patterns are consistent with most other studies of human variation and history. In contrast, growth rate showed a weaker correlation in the opposite direction. Y lineage diversity and TMRCA also decrease with distance from East Africa, supporting a model of expansion with serial founder events starting from this source. A number of individual populations diverge from these general patterns, including previously-documented examples such as recent expansions of the Yoruba in Africa, Basques in Europe and Yakut in Northern Asia. However, some unexpected demographic histories were also found, including low growth rates in the Hazara and Kalash from Pakistan, and recent expansion of the Mozabites in North Africa.

Heterogeneous patterns of gene expression diversification in mammalian gene duplicates

Farre, D., Alba, M.M. — Mon, 12 Oct 2009 08:03:09 PDT

Gene duplication is a major mechanism for molecular evolutionary innovation. Young gene duplicates typically exhibit elevated rates of protein evolution and, according to a number of recent studies, increased expression divergence. However, the nature of these changes is still poorly understood. To gain novel insights into the functional consequences of gene duplication we have undertaken an in-depth analysis of a large dataset of gene families containing primate and/or rodent-specific gene duplicates. We have found a clear tendency towards an increase in protein, promoter and expression divergence with increasing number of duplication events undergone by each gene since the human-mouse split. In addition, gene duplication is significantly associated with a reduction in expression breadth and intensity. Interestingly, it is possible to identify three main groups regarding the evolution of gene expression following gene duplication. The first group, which comprises around 25% of the families, shows patterns compatible with tissue expression partitioning. The second and largest group, comprising 33-53% of the families, shows broad expression of one of the gene copies and reduced, overlapping, expression of the other copy or copies. This can be attributed, in most cases, to loss of expression in several tissues of one or more gene copies. Finally, a substantial number of families, 19-35%, maintain a very high level of tissues expression overlap (>0.8) after tens of millions of years of evolution. These families may have been subject to selection for increased gene dosage.

Estimates of the effect of natural selection on protein coding content

Yap, V. B., Lindsay, H., Easteal, S., Huttley, G. — Thu, 08 Oct 2009 10:57:27 PDT

Analysis of natural selection is key to understanding many core biological processes, including the emergence of competition, co-operation, and complexity, and has important applications in the targeted development of vaccines. Selection is hard to observe directly but can be inferred from molecular sequence variation. For protein-coding nucleotide sequences, the ratio of nonsynonymous to synonymous substitutions () distinguishes neutrally evolving sequences ( = 1) from those subjected to purifying ( < 1) or positive Darwinian ( > 1) selection. We show that current models used to estimate are substantially biased by naturally occurring sequence compositions. We present a novel model that weights substitutions by conditional nucleotide frequencies and which escapes these artefacts. Applying it to the genomes of pathogens causing malaria, leprosy, tuberculosis and Lyme disease gave significant discrepancies in estimates with ~ 10-30% of genes affected. Our work has substantial implications for how vaccine targets are chosen and for studying the molecular basis of adaptive evolution.

The Cytophaga hutchinsonii ChTPSP: first characterized bifunctional TPS-TPP protein as putative ancestor of all eukaryotic trehalose biosynthesis proteins

Avonce, N., Wuyts, J., Verschooten, K., Vandesteene, L., Van Dijck, P. — Wed, 07 Oct 2009 09:39:00 PDT

The most widely distributed pathway to synthesize trehalose in nature consists of two consecutive enzymatic reactions with a trehalose-6-P (T6P)-synthase (TPS) enzyme, producing the intermediate T6P, and a T6P-phosphatase (TPP) enzyme, which dephosphorylates T6P to produce trehalose and inorganic phosphate. In plants these enzymes are called Class I and Class II proteins respectively with some class I proteins being active enzymes. The Class II proteins possess both TPS and TPP consensus regions, but appear to have lost enzymatic activity during evolution. Plants also contain an extra group of enzymes of small protein size of which some members have been characterized as functional TPPs. These Class III proteins have less sequence similarity with the Class I and Class II proteins. Here, we characterize for the first time, by using biochemical analysis and yeast growth complementation assays, the existence of a natural TPS-TPP bifunctional enzyme, found in the bacterial species Cytophaga hutchinsonii. Through phylogenetic analysis we show that prokaryotic genes such as ChTPSP might be the ancestor of the eukaryotic trehalose biosynthesis genes. Second, we show that plants have recruited during evolution, possibly by horizontal transfer from bacteria such as Rhodopherax ferrireducens, a new type of small protein, encoding TPP activity, which have been named Class III proteins. RfTPP has very high TPP activity upon expression in yeast. Finally, we demonstrate that TPS gene duplication, the recruitment of the class III enzymes and recruitment of a N-terminal regulatory element, that regulates the class I enzyme activity in higher plants, were initiated very early in eukaryan evolution as the three classes of trehalose biosynthesis genes are already present in the alga Ostreococcus tauri.

Inference and Characterization of Horizontally Transferred Gene Families using Stochastic Mapping

Cohen, O., Pupko, T. — Tue, 06 Oct 2009 07:30:10 PDT

Macro genomic events, in which genes are gained and lost, play a pivotal evolutionary role in microbial evolution. Nevertheless, probabilistic evolutionary models describing such events and methods for their robust inference are considerably less developed than existing methodologies for analyzing site-specific sequence evolution. Here we present a novel method for the inference of gains and losses of gene families. First, we develop probabilistic evolutionary models describing the dynamics of gene family content, which are more biologically realistic than previously suggested models. In our likelihood-based models, gains and losses are represented by transitions between presence and absence, given an underlying phylogeny. We employ a mixture-model approach in which we allow both the gain rate and the loss rate to vary among gene families. Second, we use these models together with the analytic implementation of stochastic mapping to infer branch specific events. Our novel methodology allows us to infer and quantify Horizontal Gene Transfer (HGT) events. This enables us to rank various gene families and lineages according to their propensity to undergo gains and losses. Applying our methodology to 4,873 gene families shows that (1) the novel mixture models describe the observed variability in gene family content among microbes significantly better than previous models; (2) The stochastic mapping approach enables accurate inference of gain and loss events based on simulations; (3) At least 34% of the gene families analyzed are inferred to have experienced HGT at least once during their evolution; (4) Gene families that were inferred to experience HGT are both enriched and depleted with respect to specific functional categories.

Infrequent transitions between saline and fresh waters in one of the most abundant microbial lineages (SAR11)

Logares, R., Brate, J., Heinrich, F., Shalchian-Tabrizi, K., Bertilsson, S. — Tue, 06 Oct 2009 07:30:09 PDT

The aquatic bacterial group SAR11 is one of the most abundant organisms on Earth, with an estimated global population size of 2.4 x 10²⁸ cells in the oceans. Members of SAR11 have also been detected in brackish and fresh waters, but the evolutionary relationships between the species present in the different environments have been ambiguous. In particular, it was not clear how frequently this lineage has crossed the saline-freshwater boundary during its evolutionary diversification. Due to the huge population size of SAR11 and the potential of microbes for long-distance dispersal, we hypothesized that environmental transitions could have occurred repeatedly during the evolutionary diversification of this group. Here, we have constructed extensive 16S rDNA-based molecular phylogenies and undertaken metagenomic data analyses to assess the frequency of saline-freshwater transitions in SAR11 and to investigate the evolutionary implications of this process. Our analyses indicated that very few saline-freshwater transitions occurred during the evolutionary diversification of SAR11, generating genetically distinct saline and freshwater lineages that do not appear to exchange genes extensively via horizontal gene transfer. In contrast to lineages from saline environments, extant freshwater taxa from diverse, and sometimes distant, geographic locations were very closely related. This points to a rapid diversification and dispersal in fresh waters or to slower evolutionary rates in freshwater SAR11 when compared to marine counterparts. In addition, the colonization of both saline and fresh waters appears to have occurred early in the evolution of SAR11. We conclude that the different biogeochemical conditions that prevail in saline and fresh waters have likely prevented the environmental transitions in SAR11, promoting the evolution of clearly distinct lineages in each environment.

Footprints of X-to-Y gene conversion in recent human evolution

Trombetta, B., Cruciani, F., Underhill, P. A., Sellitto, D., Scozzari, R. — Tue, 06 Oct 2009 19:43:10 PDT

Different X-homologous regions of the male specific portion of the human Y chromosome (MSY) are characterized by a different content of putative SNPs, as reported in public databases. The possible role of X-to-Y non allelic gene conversion in contributing to these differences remains poorly understood. We explored this issue by analyzing sequence variation in three regions of the MSY characterized by a different degree of X-Y similarity and a different density of putative SNPs: the PCDH11Y gene in the X-transposed (X-Y identity 99%, high putative SNP content); the TBL1Y gene in the X-degenerate (X-Y identity 86%-88%, low putative SNP content) and VCY genes-containing region in the P8 palindrome (X-Y identity 95%, low putative SNP content). Present findings do not provide any evidence for gene conversion in the PCDH11Y and TBL1Y genes; they also strongly suggest that most putative SNPs of the PCDH11Y gene (and possibly the entire X-transposed region) are most likely X-Y paralogous sequence variants, which have been entered in the databases as SNPs. On the other hand, clear evidence for the VCY genes in the P8 palindrome having acted as an acceptor of X-to-Y gene conversion was obtained. A rate of 1.8 x 10^-7 X-to-Y conversions/bp/year was estimated for these genes. These findings indicate that in the VCY region of the MSY, X-to-Y gene conversion can be highly effective to increase the level of diversity among human Y chromosomes, and suggest an additional explanation for the ability of the Y chromosome to retard degradation during evolution. Present data are expected to pave the way for future investigations on the role of non-allelic gene conversion in double strand break repair and the maintenance of Y chromosome integrity.

A statistical evaluation of models for the initial settlement of the American continent emphasizes the importance of gene flow with Asia

Mon, 05 Oct 2009 00:41:45 PDT

While there is agreement in that the Bering Strait was the entry point for the initial colonization of the American continent, there is considerable uncertainty regarding the timing and pattern of human migration from Asia to America. In order to perform a statistical assessment of the relative probability of alternative migration scenarios and to estimate key demographic parameters associated with them, we used an Approximate Bayesian Computation (ABC) framework to analyze a dataset of 401 autosomal microsatellite loci typed in 29 Native American populations. A major finding is that a single, discrete, wave of colonization is highly inconsistent with observed levels of genetic diversity. A scenario with two discrete migration waves is also not supported by the data. The current genetic diversity of Amerindian populations is best explained by a third model involving recurrent gene flow between Asia and America, after initial colonization. We estimate that this colonization involved about 100 individuals and occurred some 13,000 years ago; in agreement with well established archeological data.

Phylogenetic distributions and histories of proteins involved in anaerobic pyruvate metabolism in eukaryotes

Hug, L. A., Stechmann, A., Roger, A. J. — Mon, 05 Oct 2009 00:41:45 PDT

Protists that live in low oxygen conditions often oxidize pyruvate to acetate via anaerobic ATP-generating pathways. Key enzymes that commonly occur in these pathways are pyruvate:ferredoxin oxidoreductase (PFO) and [FeFe]-hydrogenase (H₂ase) as well as the associated [FeFe]-H₂ase maturase proteins HydE, HydF, and HydG. Determining the origins of these proteins in eukaryotes is of key importance to understanding the origins of anaerobic energy metabolism in microbial eukaryotes. We conducted a comprehensive search for genes encoding these proteins in available whole genomes and expressed sequence tag (EST) data from diverse eukaryotes. Our analyses of the presence/absence of eukaryotic PFO, [FeFe]-H₂ase, and H₂ase maturase sequences across eukaryotic diversity reveals orthologs of these proteins encoded in the genomes of a variety of protists previously not known to contain them. Our phylogenetic analyses revealed: (i) extensive lateral gene transfers of both PFO and [FeFe]-H₂ase in eubacteria, (ii) decreased support for the monophyly of eukaryote PFO domains, and, (iii) that eukaryotic [FeFe]-H₂ases are not monophyletic. Although there are few eukaryote [FeFe]-H₂ase maturase orthologs characterized, phylogenies of these proteins do recover eukaryote monophyly, although a consistent eubacterial sister group for eukaryotic homologs could not be determined. An exhaustive search for these five genes in diverse genomes from two representative eubacterial groups, the Clostridiales and the -Proteobacteria, shows that while these enzymes are nearly universally present within the former group, they are very rare in the latter. No -proteobacterial genome sequenced to date encodes all five proteins. Molecular phylogenies and the extremely restricted distribution of PFO, [FeFe]-H₂ases and their associated maturases within the -Proteobacteria do not support a mitochondrial origin for these enzymes in eukaryotes. However, the unexpected prevalence of PFO, pyruvate:NADP oxidoreductase, [FeFe]-H₂ase, and the maturase in genomes of diverse eukaryotes indicates these enzymes have an important role in the evolution of microbial eukaryote energy metabolism.

Isoform Divergence of the Filamin Family of Proteins

Kesner, B. A., Milgram, S. L., Temple, B. R. S., Dokholyan, N. V. — Mon, 05 Oct 2009 00:41:44 PDT

The vertebrate filamin family (A, B and C) is part of the spectrin family of actin cross-linking proteins. Family members share high sequence similarity (>64%) and have both common and isoform distinct functionality. To identify the basis for isoform specific functionality, we perform an evolutionary trace of chordate filamin at the granularity of single residues. Our trace methodology is constrained to focus on neo-functionality by requiring that one isoform remain the ancestral type while at least one isoform has an accepted mutation. We call divergence meeting these characteristics "class-distinctive." To obtain a temporal and spatial context for class-distinctive residues, we derive an all-atom model of full-length filamin A by homology modeling and joining individual domains. We map onto our model both conserved and class-distinctive residues along with the period (Teleostei, Amphibian, Mammalian) in which they diverged. Our phylogenetic analysis suggests that filamins diverged from a common ancestral gene between urochordate and vertebrate lineages. Filamins also diverged the most just after gene duplication, in the Teleostei period, with filamin C remaining closest to ancestral filamin. At the residue level, domains with well-characterized interfaces, IgFLN 17 and IgFLN 21, have diverged in potentially critical residues in their adhesion protein binding interfaces, signifying that isoforms may bind or regulate ligand binding differentially. Similarly, isoform divergence in a region associated with F-actin binding regulation suggests that isoforms differentially regulate F-actin binding. In addition, we observe some class-distinctive residues in the vicinity of missense mutations that cause filamin A and B associated skeletal disorders. Our analysis, utilizing both spatial and temporal granularity, has identified potentially important residues responsible for vertebrate filamin isoform specific divergence – significantly in regions where few binding partners have been discovered to date – and suggests yet to be discovered filamin binding partners, and isoform specific differential regulation with these binding partners.

Chordate Hox and ParaHox Gene Clusters Differ Dramatically in Their Repetitive Element Content

Osborne, P. W., Ferrier, D. E. K. — Mon, 05 Oct 2009 00:41:43 PDT

The ParaHox and Hox gene clusters control aspects of animal anterior-posterior development, and are related as paralogous evolutionary sisters. Despite this relationship it is not clear if the clusters operate in similar ways, with similar constraints. To compare clusters we examined the transposable element (TE) content of amphioxus and mammalian ParaHox and Hox clusters. Chordate Hox clusters are known to be largely devoid of TEs, possibly due to gene regulation and constraints on clustering in these animals. Here we describe several novel amphioxus TEs and show the amphioxus ParaHox cluster is a hotspot for TE insertion. TE contents of mammalian ParaHox loci are at background levels, in stark contrast to chordate Hox clusters. This marks a significant difference between Hox and ParaHox clusters. The presence of so many potentially disruptive elements implies selection constrains these ParaHox clusters as they have not dispersed despite 500 MY of evolution for each lineage.

Hyper-variability of ascidian mitochondrial gene order: exposing the myth of deuterostome organelle genome stability

Gissi, C., Pesole, G., Mastrototaro, F., Iannelli, F., Guida, V., Griggio, F. — Thu, 01 Oct 2009 04:47:36 PDT

The few sequenced mitochondrial (mt) genomes of the class Ascidiacea (Chordata, Tunicata), mostly belonging to congeneric species of the Phlebobranchia order, show extraordinary gene order rearrangements. In order to assess if this hypervariability in gene order is a general feature of Ascidiacea, we here report the gene arrangement of five ascidians belonging to the Aplousobranchia and Stolidobranchia orders. Our data show that Ascidiacea are characterized by: (1) extensive gene order rearrangements both within and between the three major lineages; (2) lack of significant similarities to the gene order of other deuterostomes; and (3) an extent of rearrangements comparable to that of Mollusca (especially the Gastropoda, Bivalvia and Scaphopoda classes), a phylum with highly-rearranged mtDNAs. The only conserved feature is the location of all genes on the same strand, which suggests that selective constrains are related to the mt transcription. Finally, a higher mobility of the tRNA genes is undetectable because of saturation effect, and only the partially conserved cox2-cob gene block seems to retain some phylogenetic signals.

How robust are "Isolation with Migration" analyses to violations of the IM model? A simulation study

Strasburg, J. L., Rieseberg, L. H. — Wed, 30 Sep 2009 06:03:11 PDT

Methods developed over the past decade have made it possible to estimate molecular demographic parameters such as effective population size, divergence time, and gene flow with unprecedented accuracy and precision. However, they make simplifying assumptions about certain aspects of the species’ histories and the nature of the genetic data, and it is not clear how robust they are to violations of these assumptions. Here we use simulated datasets to examine the effects of a number of violations of the "Isolation with Migration" (IM) model, including intralocus recombination, population structure, gene flow from an unsampled species, linkage among loci, and divergent selection, on demographic parameter estimates made using the program IMa. We also examine the effect of having data that fits a nucleotide substitution model other than the two relatively simple models available in IMa. We find that IMa estimates are generally quite robust to small to moderate violations of the IM model assumptions, comparable to what is often encountered in real-world scenarios. In particular, population structure within species, a condition encountered to some degree in virtually all species, has little effect on parameter estimates even for fairly high levels of structure. Likewise, most parameter estimates are robust to significant levels of recombination when datasets are pared down to apparently non-recombining blocks, although substantial bias is introduced to several estimates when the entire dataset with recombination is included. In contrast, a poor fit to the nucleotide substitution model can result in an increased error rate, in some cases due to a predictable bias and in other cases due to an increase in variance in parameter estimates among datasets simulated under the same conditions.

Retracing evolution of red fluorescence in GFP-like proteins from Faviina corals

Field, S. F., Matz, M. V. — Wed, 30 Sep 2009 06:03:11 PDT

GFP-like fluorescent proteins represent is a convenient experimental model to study evolution of novelty at the molecular level. Here we focus on the origin of Kaede-like red fluorescent proteins characteristic of the corals of the Faviina suborder. We demonstrate, using an original approach involving resurrection and analysis of the library of possible evolutionary intermediates, that it takes on the order of 12 mutations, some of which strongly interact epistatically, to fully recapitulate the evolution of a red fluorescent phenotype from the ancestral green. Five of the identified mutations would not have been found without the help of ancestral reconstruction, since the corresponding site states are shared between extant red and green proteins due to their recent descent from a dual-function common ancestor. Seven of the 12 mutations are affecting residues that are not in close contact with the chromophore and thus must exert their effect indirectly through adjustments of the overall protein fold; the relevance of these mutations could not have been anticipated from the purely theoretical analysis of the protein's structure. Our results introduce a powerful experimental approach for comparative analysis of functional specificity in protein families even in the cases of pronounced epistasis, provide foundation for the detailed studies of evolutionary trajectories leading to novelty and complexity, and will help rational modification of existing fluorescent labels.

Nucleotide polymorphism at a gene (Pgi) under balancing selection in a butterfly metapopulation

Wheat, C. W., Haag, C. R., Marden, J. H., Hanski, I., Frilander, M. J. — Wed, 30 Sep 2009 06:03:10 PDT

The Glanville fritillary butterfly (Melitaea cinxia, Nymphalidae) has a large, well-studied metapopulation in the Åland Islands in Finland. Previous studies have found that the common allozyme genotypes at the phosphoglucose isomerase (PGI) locus are associated with individual variation in performance and fitness, with phenotypic data suggesting ongoing balancing selection via heterozygote advantage. Here we analyse nucleotide polymorphism in the coding region of the Pgi gene. Pgi is exceptionally polymorphic, in contrast to three other metabolic genes (Mdh, Idh and Gapdh) with low levels of polymorphism. Most of the variation is due to two common haplotype clades, which are highly divergent and exhibit extensive linkage disequilibrium. These two clades correspond to the two most common allozyme alleles previously studied. Molecular tests of selection and coalescence simulations indicate that patterns of nucleotide polymorphism depart from neutrality and are consistent with long-term balancing selection. The split between the two main haplotype clades is estimated to predate the last common ancestor of a clade of five extant Melitaea species. Comparative structural analysis of Pgi polymorphism in M. cinxia and the unrelated Colias eurytheme butterfly suggests a similar but not identical target of balancing selection. Our results indicate convergent evolution between these two species at both the phenotypic and molecular level.

Rapid likelihood analysis on large phylogenies using partial sampling of substitution histories

Jason de Koning, A. P., Gu, W., Pollock, D. D. — Fri, 25 Sep 2009 06:38:05 PDT

Likelihood-based approaches can reconstruct evolutionary processes in greater detail and with better precision from larger datasets. The extremely large comparative genomic datasets that are now being generated thus create new opportunities for understanding molecular evolution, but analysis of such large quantities of data poses escalating computational challenges. Recently developed Markov chain Monte Carlo (MCMC) methods that augment substitution histories are a promising approach to alleviate these computational costs. We analysed the computational costs of several such approaches, considering how they scale with model and dataset complexity. This provided a theoretical framework to understand the most important computational bottlenecks, leading us to combine novel variations of our conditional pathway integration approach with recent advances made by others. The resulting technique (‘partial sampling’ of substitution histories) is considerably faster than all other approaches we considered. It is accurate, simple to implement, and scales exceptionally well with dimensions of model complexity and dataset size. In particular, the time complexity of sampling unobserved substitution histories using the new method is much faster than previously existing methods, and model parameter and branch length updates are independent of dataset size. We compared the performance of methods on a 224-taxon set of mammalian cytochrome b sequences. For a simple nucleotide substitution model, partial sampling was at least 10 times faster than the PhyloBayes program, which samples substitutions in continuous time, and about 100 times faster than when using fully integrated substitution histories. Under a general reversible model of amino-acid substitution, the partial sampling method was 1600 times faster than when using fully integrated substitution histories, confirming significantly improved scaling with model state-space complexity. Partial sampling of substitutions thus dramatically improves the utility of likelihood approaches for analysing complex evolutionary processes on large datasets.

A genome sequence survey for Ascogregarina taiwanensis supports evolutionary affiliation, but metabolic diversity between a gregarine and Cryptosporidium

Thu, 24 Sep 2009 05:46:57 PDT

We have performed a whole genome sequence survey (GSS) for the gregarine, Ascogregarina taiwanensis, and herein describe features both unique to this early diverging apicomplexan and properties that unite it with Cryptosporidium, the Coccidia and the Apicomplexa. Phylogenetic trees inferred from a concatenated protein sequence comprised of 10,750 amino acid positions, as well as the large subunit rRNA genes, robustly support phylogenetic affinity of Ascogregarina with Cryptosporidium at the base of the apicomplexan clade. Unlike Cryptosporidium, Ascogregarina possesses numerous mitochondrion-associated pathways and proteins, including enzymes within the Krebs cycle and a cytochrome-based respiratory chain. Ascogregarina further differs in a capacity for de novo synthesis of pyrimidines and amino acids. Ascogregarina shares with Cryptosporidium a Type I fatty acid synthase, and likely a polyketide synthase. Cryptosporidium and Ascogregarina possess a large repertoire of multi-domain surface proteins that align it with Toxoplasma and are proposed to be involved in coccidian-like functions. Four families of retrotransposable elements were identified, and thus retroelements are present in Ascogregarina and Eimeria but not in other apicomplexans that have been analyzed. The sum observations suggest that Ascogregarina and Cryptosporidium share numerous molecular similarities, including coccidian-like features to the exclusion of Haemosporidia and Piroplasmida, but also differ from each other significantly in their metabolic capacity.

The evolution of amastin surface glycoproteins in trypanosomatid parasites

Jackson, A. P. — Thu, 24 Sep 2009 02:02:00 PDT

Amastin is a transmembrane glycoprotein found on the cell surfaces of trypanosomatid parasites. Encoded by a large, diverse gene family, amastin was initially described from the intracellular, amastigote stage of Trypanosoma cruzi and Leishmania donovani. Genome sequences have subsequently shown that the amastin repertoire is much larger in Leishmania relative to Trypanosoma. However, it is not known when this expansion occurred, whether it is associated with the origins of Leishmania and vertebrate parasitism itself, or prior to this. To examine the timing of amastin diversification, as well as the evolutionary mechanisms regulating gene repertoire and sequence diversity, this study sequenced the genomic regions containing amastin loci from two related insect parasites (Leptomonas seymouri and Crithidia sp.), and estimated a phylogeny for these and other amastin sequences. The phylogeny shows that amastin includes four subfamilies with distinct genomic positions, secondary structures, and evolution, which were already differentiated in the ancestral trypanosomatid. Diversification in Leishmania was initiated from a single ancestral locus on chromosome 34, with rapid derivation of novel loci through transposition and accelerated sequence divergence. This is absent from related organisms showing that diversification occurred after the origin of Leishmania. These results describe a substantial elaboration of amastin repertoire directly associated with the origin of Leishmania, suggesting that some amastin genes evolved novel functions crucial to cell function in leishmanial parasites after the acquisition of a vertebrate host.

Molecular Evolution Within and Between Self-Incompatibility Specificities

Tue, 22 Sep 2009 07:48:33 PDT

Genes under multiallelic balancing selection have sharply contrasted evolutionary dynamics across time scales, with much longer coalescence time among functionally distinct allelic lines but much shorter coalescence time among gene copies within allelic lines as compared to the genomic background. In this paper, we combine theoretical and empirical approaches to investigate patterns of molecular evolution within and between self-incompatibility specificities. We first use numerical simulations to investigate coalescence times within allelic lines in a subdivided population for a sporophytic self-incompatibility system. We then report on a comprehensive analysis of nucleotide polymorphism among gene copies within five distinct allelic lines in the closely related Arabidopsis halleri and A. lyrata. In line with our model predictions, we find that the observed level of polymorphism among gene copies was generally low but differed among allelic lines. The data provides compelling direct evidence for recombination and/or gene conversion not only within the two most recessive allelic lines but also between two closely related but distinct allelic lines, suggesting that recombination at the Arabidopsis self-incompatibility locus is possible in the absence of large sequence divergence among haplotypes. We observed shared polymorphic sites between the two species in one allelic line and strikingly similar haplotypes in another allelic line. We discuss whether convergent evolution may have led to this pattern, and suggest that these observations are consistent with ongoing or very recent introgression, as previously documented.

Zebu cattle are an exclusive legacy of the South Asia Neolithic

Mon, 21 Sep 2009 07:01:00 PDT

Animal domestication was a major step forward in human prehistory, contributing to the emergence of more complex societies. At the time of the Neolithic transition, zebu cattle (Bos indicus) were probably the most abundant and important domestic livestock species in Southern Asia. Although archaeological evidence points towards the domestication of zebu cattle within the Indian subcontinent, the exact geographic origins and phylogenetic history of zebu cattle remains uncertain. Here, we report evidence from 844 zebu mitochondrial DNA (mtDNA) sequences surveyed from 19 Asiatic countries comprising eight regional groups, which identify two distinct mitochondrial haplogroups, termed I1 and I2. The marked increase in nucleotide diversity (P < 0.001) for both the I1 and I2 haplogroups within the northern part of the Indian subcontinent is consistent with an origin for all domestic zebu in this area. For haplogroup I1, genetic diversity was highest within the Indus Valley among the three hypothesized domestication centers (Indus Valley, Ganges and South India). These data support the Indus Valley as the most likely center of origin for the I1 haplogroup and a primary center of zebu domestication. However, for the I2 haplogroup a complex pattern of diversity is detected, preventing the unambiguous pinpointing of the exact place of origin for this zebu maternal lineage. Our findings are discussed with respect to the archaeological record for zebu domestication within the Indian subcontinent.

Novel Internal Regions of Fluorescent Proteins Undergo Divergent Evolutionary Patterns

Mon, 21 Sep 2009 07:00:59 PDT

Over the past decade, Fluorescent Proteins (FPs) have become ubiquitous tools in biological research. Yet little is known about the natural function or evolution of this superfamily of proteins that originates from marine organisms. Using molecular phylogenetic analyses of 102 naturally-occurring Cyan Fluorescent Proteins (CFPs), Green Fluorescent Proteins (GFPs), Red Fluorescent Proteins (RFPs) as well the non-fluorescent (purple-blue) protein sequences (including new FPs from Lizard Island, Australia) derived from organisms with known geographic origin, we show that FPs consist of two distinct and novel regions that have evolved under opposite and sharply divergent evolutionary pressures. A central region is highly conserved, and while it contains the residues that form the chromophore, its evolution does not track with fluorescent color and evolves independently from the rest of the protein. By contrast, the regions enclosing this central region are under strong positive selection pressure to vary its sequence, and yet segregate well with fluorescence color emission. We did not find a significant correlation between geographic location of the organism from which the FP was isolated and molecular evolution of the protein. These results define for the first time, two distinct regions based on evolution, for this highly compact protein. The findings have implications for more sophisticated bioengineering of this molecule as well as studies directed towards understanding the natural function of FPs.

An ancient fission of mitochondrial cox1

Gawryluk, R. M.R., Gray, M. W. — Fri, 18 Sep 2009 07:12:02 PDT

Many genes inherited from the -proteobacterial ancestor of mitochondria have undergone evolutionary transfer to the nuclear genome in eukaryotes. In some rare cases, genes have been functionally transferred in pieces, resulting in ‘split’ proteins that presumably interact in trans within mitochondria, fulfilling the same role as the ancestral, intact protein. We describe a nucleus-encoded mitochondrial protein (here named Cox1-c) in the amoeboid protist Acanthamoeba castellanii that is homologous to the C-terminal portion of conventional mitochondrial Cox1, while the corresponding portion of the mitochondrion-encoded A. castellanii Cox1 is absent. Bioinformatics searches retrieved nucleus-encoded Cox1-c homologs in most major eukaryotic supergroups; in these cases, also, the mitochondrion-encoded Cox1 lacks the corresponding C-terminal motif. These data constitute the first report of functional re-location of a portion of cox1 to the nucleus. This transfer event was likely ancient, with the resulting nuclear cox1-c being differentially activated across the eukaryotic domain.

ZP Domain Proteins in the Abalone Egg Coat include a Paralog of VERL Under Positive Selection that Binds Lysin and 18-kDa Sperm Proteins

Aagaard, J. E., Vacquier, V. D., MacCoss, M. J., Swanson, W. J. — Fri, 18 Sep 2009 07:12:02 PDT

Identifying fertilization molecules is key to our understanding of reproductive biology, yet only a few examples of interacting sperm and egg proteins are known. One of the best characterized comes from the invertebrate archeogastropod abalone (Haliotis spp.), where sperm lysin mediates passage through the protective egg vitelline envelope (VE) by binding to the VE protein VERL. Rapid adaptive divergence of abalone lysin and VERL are an example of positive selection on interacting fertilization proteins contributing to reproductive isolation. Previously, we characterized a subset of the abalone VE proteins that share a structural feature, the ZP domain, which is common to VERL and the egg envelopes of vertebrates. Here we use additional EST sequencing and shotgun proteomics to characterize this family of proteins in the abalone egg VE. We expand 3-fold the number of known ZP domain proteins present within the VE (now 30 in total) and identify a paralog of VERL (VEZP14) that contains a putative lysin binding motif. We find that, like VERL, the divergence of VEZP14 among abalone species is driven by positive selection on the lysin binding motif alone and that these paralogous egg VE proteins bind a similar set of sperm proteins including a rapidly evolving 18-kDa paralog of lysin, which may mediate sperm egg fusion. This work identifies an egg coat paralog of VERL under positive selection and the candidate sperm proteins with which it may interact during abalone fertilization.

A timeline of the environmental genetics of the haptophytes

Liu, H., Aris-Brosou, S., Probert, I., de Vargas, C. — Wed, 16 Sep 2009 23:06:23 PDT

The use of genomic data and the rise of phylogenomics have radically changed our view of the eukaryotic tree of life at a high taxonomic level by identifying four to six "supergroups". Yet our understanding of the evolution of key innovations within each of these supergroups is limited because of poor species sampling relative to the massive diversity encompassed by each supergroup. Here we apply a multigene approach that incorporates a wide taxonomic diversity to infer the timeline of the emergence of strategic evolutionary transitions in the haptophytes, a group of ecologically and biogeochemically significant marine protists that belong to the Chromalveolata supergroup. Four genes (SSU, LSU, tufA and rbcL) were extensively analyzed under several Bayesian models to assess the robustness of the phylogeny, particularly with respect to (i) data partitioning, (ii) the origin of the genes (host vs. endosymbiont), (iii) across-site rate variation and (iv) across-lineage rate variation. We show with a relaxed clock analysis that the origin of haptophytes dates back to 824 MYA (95% highest probability density 1031-637 MYA). Our dating results show that the ability to calcify evolved earlier than previously thought, between 329-291 MYA, in the Carboniferous period, and that the transition from mixotrophy to autotrophy occurred during the same time period. Although these two transitions precede a habitat change of major diversities from coastal / neritic waters to the pelagic realm (291-243 MYA, around the P/Tr boundary event), the emergence of calcification, full autotrophy and oceanic lifestyle seem mutually independent.

Uncovering the footprint of positive selection on the X chromosome of Drosophila melanogaster

Orengo, D. J., Aguade, M. — Wed, 16 Sep 2009 08:49:49 PDT

A usual approach to detect the spatial footprint left by recent adaptive events has been to follow up putative candidates emerging from multilocus scans of variation by sequencing additional fragments. We have used a similar experimental and analytical approach to study variation at 15 independently evolving and randomly chosen regions of the X chromosome of Drosophila melanogaster. These incompletely sequenced regions, extending each over ~40 kb, were subjected to two tests of positive selection that take into account the spatial distribution of nucleotide variation. Our analysis of variation at these genomic regions in a European population of D. melanogaster has allowed us to uncover a candidate region for positive selection and to empirically evaluate the comparative performance of the two tests of selection under a bottleneck scenario. Moreover, the boundaries here estimated for both the rate of adaptive substitution () and the average selection coefficient (s) would support previous estimates obtained by maximum likelihood that suggest rather strong but uncommon positive selection.

Distributions of selectively constrained sites and deleterious mutation rates in the hominid and murid genomes

Eory, L., Halligan, D. L., Keightley, P. D. — Wed, 16 Sep 2009 08:49:48 PDT

Protein-coding sequences make up only about 1% of the mammalian genome. Much of the remaining 99% has been long assumed to be junk DNA, with little or no functional significance. Here we show that in hominids, a group with historically low effective population sizes, all classes of non-coding DNA evolve more slowly than ancestral transposable elements, and so appear to be subject to significant evolutionary constraints. Under the nearly neutral theory, we expected to see lower levels of selective constraints on most sequence types in hominids than murids, a group that is thought to have a higher effective population size. We found that this is the case for many sequence types examined, the most extreme example being 5’ UTRs, for which constraint in hominids is only about one-third that of murids. Surprisingly, however, we observed higher constraints for some sequence types in hominids, notably four-fold sites, where constraint is more than twice as high as in murids. This implies that more than about one-fifth of mutations at four-fold sites are effectively selected against in hominids. The higher constraint at four-fold sites in hominids suggests a more complex protein-coding gene structure than murids, and indicates that methods for detecting selection on protein coding sequences (e.g., using the d_N /d_S ratio), with four-fold sites as a neutral standard, may lead to biased estimates, particularly in hominids. Our constraint estimates imply that 5.4% of nucleotide sites in the human genome are subject to effective negative selection, and that there are three times as many constrained sites within non-coding sequences as within protein-coding sequences. Including coding and non-coding sites, we estimate that the genomic deleterious mutation rate U = 4.2. The mutational load predicted under a multiplicative model is therefore about 99% in hominids.

Evolutionary dynamics of rhodopsin type 2 (RH2) opsins in vertebrates

Yokoyama, S., Tada, T. — Wed, 16 Sep 2009 08:49:47 PDT

Among the five groups of visual pigments in vertebrates, the rhodopsin type 2 (RH2) group shows the largest number of gene duplication events. We have isolated three intact and one nonfunctional RH2 opsin genes each from Northern lampfish (Stenobrachius leucopsarus) and scabbardfish (Lepidopus fitchi). Using the deduced amino acid sequences of these and other representative RH2 opsin genes in vertebrates, we have estimated the divergence times and evolutionary rates of amino acid substitution at various stages of RH2 opsin evolution. The results show that the duplications of the lampfish and scabbardfish RH2 opsins have occurred ~60 and ~30 MYA, respectively. The evolutionary rates of RH2 opsins in the early vertebrate ancestors were ~0.25 x 10^-9/site/year, which increased to ~1-3 x 10^-9/site/year in euteleost lineages and to ~0.3-0.5 x 10^-9/site/year in coelacanth and tetrapods.

Origin and evolution of the unique tetra-domain hemoglobin from the hydrothermal vent scale-worm Branchipolynoe

Tue, 15 Sep 2009 11:40:09 PDT

Hemoglobin is the most common respiratory pigment in annelids. They can be intra- or extra-cellular, and this latter type can form large multimeric complexes. The hydrothermal vent scale-worms Branchipolynoe symmytilida and B. seepensis express an extracellular tetra-domain hemoglobin that is unique in annelids. We sequenced the gene for the single-domain and tetra-domain globins in these two species. The single-domain gene codes for a mature protein of 137 amino acids, and the tetra-domain gene codes for a mature protein of 552 amino acids. The single-domain gene has a typical 3 exons/2 introns structure, with introns located at their typical positions (B12.2 and G7.0). This structure is repeated four times in the tetra-domain gene, with no bridge introns or linker sequences between domains. The phylogenetic position of Branchipolynoe globins among known annelid globins revealed that, although extracellular, they cluster within the annelid intracellular globins clade, suggesting that the extracellular state of these hemoglobins is the result of convergent evolution. The tetra-domain structure likely resulted from two tandem duplications, domain 1 giving rise to domain 2 and after this the two-domain gene duplicated to produce domains 3 and 4. The high O₂ affinity of Branchipolynoe extracellular globins may be explained by the two key residues (B10Y and E7Q) in the heme pocket in each of the domains of the single and tetra-domain globins, which have been shown to be essential in the oxygen-avid hemoglobin from the nematode Ascaris suum. This peculiar globin evolutionary path seems to be very different from other annelid extracellular globins and is most likely the product of evolutionary tinkering associated with the strong selective pressure to adapt to chronic hypoxia that characterizes hydrothermal vents.

Philippine mitochondrial DNA diversity: a populated viaduct between Taiwan and Indonesia?

Tue, 15 Sep 2009 11:40:08 PDT

Relatively little is known about the genetic diversity of the Philippine population, and this is an important gap in our understanding of Southeast Asian and Oceanic prehistory. Here we describe mtDNA variation in 423 Philippine samples and analyze them in the context of the genetic diversity of other Southeast Asian populations. The majority of Philippine mtDNA types are shared with Taiwanese aboriginal groups and belong to haplogroups of post-glacial and pre-Neolithic origin which have previously been identified in East Asian and Island Southeast Asian populations. Analysis of Hypervariable Segment I sequence variation within individual mtDNA haplogroups indicates a general decrease in the diversity of the most frequent types (B4a1a, E1a1a, M7c3c) from the Taiwanese aborigines to the Philippines and Sulawesi, although calculated standard error measures overlap for these populations. This finding, together with the geographical distribution of ancestral and derived haplotypes of the B4a1a sub-clade including the Polynesian Motif, is consistent with southward dispersal of these lineages "Out of Taiwan" via the Philippines to Near Oceania and Polynesia. In addition to the mtDNA components shared with Taiwanese aborigines, complete sequence analyses revealed a minority of lineages in the Philippines which share their origins - possibly dating back to the Paleolithic - with haplogroups from Indonesia and New Guinea. Other rare lineages in the Philippines have no closely related types yet identified elsewhere.

Adaptive evolution of digestive RNASE1 genes in leaf-eating monkeys revisited: new insights from 10 additional Colobines

Yu, L., Wang, X.-y., Jin, W., Luan, P.-t., Ting, N., Zhang, Y.-p. — Fri, 11 Sep 2009 03:03:04 PDT

Pancreatic RNase genes implicated in the adaptation of the colobine monkeys to leaf-eating have long intrigued evolutionary biologists since the identification of a duplicated RNASE1 gene with enhanced digestive efficiencies in Pygathrix nemaeus. The recent emergence of two contrasting hypotheses, i.e., independent duplication and one duplication event hypotheses, make it into focus again. Current understanding of Colobine RNASE1 gene evolution largely depends on the analyses of few colobine species. The present study with more intensive taxonomic and character sampling not only provides a clearer picture of Colobine RNASE1 gene evolution, but also allows to have a more thorough understanding about the molecular basis underlying the adaptation of Colobinae to the unique leaf-feeding lifestyle. The present broader and detailed phylogenetic analyses yielded two important findings: (1) All trees based on the analyses of coding, noncoding and both regions provided consistent evidence indicating RNASE1 duplication occurred after Asian and African colobines speciation, i.e., independent duplication hypothesis; (2) No obvious evidence of gene conversion in RNASE1 gene was found, favoring independent evolution of Colobine RNASE1 gene duplicates. The conclusion drawn from previous studies that gene conversion has played a significant role in the evolution of Colobine RNASE1 was not supported. Our selective constraint analyses also provided interesting insights, with significant evidence of positive selection detected on ancestor lineages leading to duplicated gene copies. The identification of a handful of new adaptive sites and amino acid changes that have not been characterized previously also provide a necessary foundation for further experimental investigations of RNASE1 functional evolution in Colobinae.

Analysis of the immunoglobulin light chain genes in zebra finch: evolutionary implications

Das, S., Mohamedy, U., Hirano, M., Nei, M., Nikolaidis, N. — Thu, 10 Sep 2009 05:43:57 PDT

All jawed vertebrates produce immunoglobulins (IGs) as a defense mechanism against pathogens. Typically, immunoglobulins are composed of two identical heavy chains (IGH) and two identical light chains (IGL). Most tetrapod species encode more than one isotype of light chains. Chicken is the only representative of birds for which genomic information is currently available and is an exception to the above rule, because it encodes only a single IGL isotype (i.e., lambda). Here, we show that the genome of zebra finch, another bird species, encodes a single IGL isotype, i.e., lambda, like the chicken. These results strongly suggest that the second isotype (i.e., kappa) present in both reptiles and mammals was lost in a very early stage of bird evolution. Furthermore, we show that both chicken and zebra finch contain a single set of functional variable, joining, and constant region genes and multiple variable region pseudogenes. The latter finding suggests that this type of genomic organization was already present in the common ancestor of these bird species and remained unchanged over a long evolutionary time. This conservation is in contrast with the high levels of variation observed in the mammalian IGL loci. The presence of a single functional variable region gene followed by multiple variable pseudogenes in zebra finch suggest that this species may be generating antibody diversity by a gene conversion-like mechanism like the chicken.

Selection to maintain amino acid differences between paralogous heat shock protein genes in yeast

Takuno, S., Innan, H. — Thu, 10 Sep 2009 05:43:57 PDT

A genome scan for the signatures of selection for paralogous functional amino acid differences were performed with yeast genomes. This recently developed method makes it possible to localize the target site(s) of selection. We found that two gene pairs have strong signatures of selection. The two pairs of duplicated genes happened to be heat shock genes (Ssa1/Ssa2 and Ssb1/Ssb2), which have similar protein structures to each other while the amino acid sequence identity between Ssa and Ssb is not high (~ 60%). Interestingly, the two gene pairs exhibit signature of selection at almost identical positions within the substrate-binding domain-β. Because this domain specifies the substrate polypeptides, it is presumed that functional divergence may be advantageous in this domain. Evolutionary analysis demonstrated that the observed divergence in the two gene pairs have been maintained in many yeast species independently, suggesting long-term operation of strong selection.

Lower linkage disequilibrium at CNVs is due to both recurrent mutation and transposing duplications

Schrider, D. R., Hahn, M. W. — Thu, 10 Sep 2009 05:43:56 PDT

Copy number variants (CNVs) within humans can have both adaptive and deleterious effects. Because of their phenotypic significance, researchers have attempted to find single nucleotide polymorphisms (SNPs) in high linkage disequilibrium with CNVs to use in genome-wide association studies. However, studies have found that CNVs are less likely to be in strong linkage disequilibrium with flanking markers. We hypothesized that this "taggability gap" can be explained by duplication events that place paralogous sequences far apart. In support of our hypothesis, we find that duplications are significantly less likely than deletions to have a "tag" SNP, even after controlling for CNV length, allele frequency, and availability of appropriate flanking SNPs. Using a novel likelihood method, we are able to show that many complex CNVs—those due to multiple duplication or deletion polymorphisms—are made up of two loci with little linkage disequilibrium between them. Additionally, we find that many polymorphic duplications detected in a recent clone-based study are located far from their parental loci. We also examine two other common hypotheses for the taggability gap, and find that recurrent mutation of both deletions and duplications appears to have an effect on linkage disequilibrium, but that lower SNP-density around CNVs has no effect. Overall, our results suggest that a substantial fraction of CNVs caused by duplication cannot be tagged by markers flanking the parental locus because they have changed genomic location.

Measuring the Sensitivity of Single-Locus "Neutrality Tests" Using a Direct Perturbation Approach

Garrigan, D., Lewontin, R., Wakeley, J. — Thu, 10 Sep 2009 05:43:56 PDT

A large number of statistical tests have been proposed to detect natural selection based on a sample of variation at a single genetic locus. These tests measure the deviation of the allelic frequency distribution observed within populations from the distribution expected under a set of assumptions that includes both neutral evolution and equilibrium population demography. The present study considers a new way to assess the statistical properties of these tests of selection, by their behavior in response to direct perturbations of the steady-state allelic frequency distribution, unconstrained by any particular non-equilibrium demographic scenario. Results from Monte Carlo computer simulations indicate that most tests of selection are more sensitive to perturbations of the allele frequency distribution that increase the variance in allele frequencies than to perturbations that decrease the variance. Simulations also demonstrate that it requires, on average, 4N generations (N is the diploid effective population size) for tests of selection to relax to their theoretical, steady state distributions following different perturbations of the allele frequency distribution to its extremes. This relatively long relaxation time highlights the fact that these tests are not robust to violations of the other the assumptions of the null model besides neutrality. Lastly, genetic variation arising under an example of a regularly cycling demographic scenario is simulated. Tests of selection performed on this last set of simulated data confirms the confounding nature of these tests for the inference of natural selection, under a demographic scenario that likely holds for many species. The utility of using empirical, genomic distributions of test statistics, instead of the theoretical steady state distribution, is discussed as an alternative for improving the statistical inference of natural selection.

Molecular phylogenetics, temporal diversification and principles of evolution in the mustard family (Brassicaceae)

Thu, 10 Sep 2009 05:43:55 PDT

Brassicaceae is an important family at both the agronomic and scientific levels. The family not only inlcudes several model species, but it is also becoming an evolutionary model at the family-level. However, resolving the phylogenetic relationships within the family has been problematic, and a large-scale molecular phylogeny in terms of generic sampling and number of genes is still lacking. In particular, the deeper relationships within the family, e.g. between the three major recognized lineages, prove particularly hard to resolve. Using a slow-evolving mitochondrial marker (nad4 intron 1), we reconstructed a comprehensive phylogeny in generic representation for the family. In addition, and because resolution was very low in previous single marker phylogenies, we adopted a supermatrix approach by concatenating all checked and reliable sequences available on GenBank as well as new sequences for a total 207 curently recognized genera and eight molecular markers representing a comprehensive coverage of all three genomes. The supermatrix was dated under an uncorrelated relaxed molecular clock using a direct fossil calibration approach. Finally, a lineage-through-time-plot (LTT) and rates of diversification for the family were generated. The resulting tree, the largest in number of genera and markers sampled to date and covering the whole family in a representative way, provides important insights into the evolution of the family on a broad scale. The backbone of the tree remained largely unresolved and is interpreted as the consequence of early rapid radiation within the family. The age of the family was inferred to be 37.6 (24.2–49.4) Ma, which largely agrees with previous studies. The ages of all major lineages and tribes are also reported. Analysis of diversification suggests that Brassicaceae underwent a rapid period of diversification, after the split with the early diverging tribe Aethionemeae. Given the dates found here, the family appears to have originated under a warm and humid climate ca. 37 Ma ago. We suggest that the rapid radiation detected was caused by a global cooling during the Oligocene coupled with a genome duplication event. This duplication could have allowed the family to rapidly adapt to the changing climate.

Lineage-specific adaptive evolution of the centromeric protein CENH3 in diploid and allotetraploid Oryza species

Hirsch, C. D., Wu, Y., Yan, H., Jiang, J. — Wed, 09 Sep 2009 05:59:13 PDT

Centromeres in eukaryotic species are defined by the presence of a centromere-specific histone H3 variant, CENH3. CENH3 plays a key role in recruiting other centromeric proteins, thus, it is the central component in kinetochore formation and centromere function. The CENH3 proteins in several plant and animal species were found to be under positive selection, which was hypothesized to respond to the rapid changing of the repetitive DNA sequences associated with the centromeres. Here we report the expression and evolution of the CenH3 genes in two allotetraploid rice species as well as their representative diploid progenitor species. Both copies of the CenH3 genes were transcribed in the two allotetraploid species and showed a non-preferential expression pattern. Contrasting positive and stabilizing selection of the CenH3 genes was associated with different diploid Oryza species. This lineage-specific adaptive evolution of CENH3 was maintained in the two allotetraploid species. Thus, we demonstrate that the allopolyploidization events did not alter the expression or evolutionary patterns of the CenH3 genes in the Oryza species.

Cryptic sex in the smallest eukaryotic marine green alga

Grimsley, N., Pequin, B., Bachy, C., Moreau, H., Piganeau, G. — Fri, 04 Sep 2009 08:02:01 PDT

Ostreococcus spp are common worldwide oceanic picoeukaryotic pelagic algae. The complete genomes of 3 strains from different ecological niches revealed them to represent biologically distinct species despite their identical cellular morphologies (cryptic species). Their tiny genomes (13Mb), with ~20 chromosomes, are colinear and densely packed with coding sequences, but no sexual life-cycle has been described. Seventeen new strains of one of these species, O. tauri, were isolated from 98 seawater samplings from the N.W. Mediterranean, by filtering, culturing, cloning and plating for single colonies, and identification by sequencing their ribosomal 18S gene. In order to find genetic markers for detection of polymorphisms and sexual recombination, we used an in silico approach to screen available genomic data. Intergenic regions of DNA likely to evolve neutrally were analysed following PCR-amplification of sequences using flanking primers from adjacent conserved coding sequences that were present as syntenic pairs in 2 different species of Ostreococcus. Analyses of such DNA regions from 8 marker loci on 2 chromosomes from each strain revealed that the isolated O. tauri clones were haploid and that the overall level of polymorphism was ~0.01. Four different genetic tests for recombination showed that sexual exchanges must be inferred to account for the between-locus and between-chromosome marker combinations observed. However, our data suggest that sexual encounters are infrequent, since we estimate the frequency of meioses/mitoses among the sampled strains to be 10^–6. O. tauri and related species encode and express core genes for mitosis and meiosis, but their mechanisms of cell division and recombination nevertheless remain enigmatic since a classical eukaryotic spindle with 40 canonical microtubules would be much too large for the available ~0.9 µm³ cellular volume.

A comprehensive classification and evolutionary analysis of plant homeobox genes

Mukherjee, K., Brocchieri, L., Burglin, T. R. — Fri, 04 Sep 2009 08:02:02 PDT

The full complement of homeobox transcription-factor sequences, including genes and pseudogenes, was determined from the analysis of ten complete genomes from flowering plants, moss, Selaginella, unicellular green algae and red algae. Our exhaustive genome-wide searches resulted in the discovery in each class of a greater number of homeobox genes than previously reported. All homeobox genes can be unambiguously classified by sequence evolutionary analysis into fourteen distinct classes also characterized by conserved intron/exon structure and by unique co-domain architectures. We identified many new genes belonging to previously defined classes (HD-ZIP I to IV, BEL, KNOX, PLINC, WOX). Other newly identified genes allowed us to characterize PHD, DDT, NDX and LD genes as members of four new evolutionary classes and to define two additional classes, which we named SAWADEE and PINTOX. Our comprehensive analysis allowed us to identify several newly characterized conserved motifs, including novel zinc-finger motifs in SAWADEE and DDT. Members of the BEL and KNOX classes were found in Chlorobionta (green plants) and in Rhodophyta. We found representatives of the DDT, WOX and PINTOX classes only in green plants, including unicellular green algae, moss and vascular plants. All fourteen homeobox gene classes were represented in flowering plants, Selaginella and moss, suggesting that they had already differentiated in the last common ancestor of moss and vascular plants.

Evolution of 7SK RNA and its Protein Partners in Metazoa

Fri, 04 Sep 2009 08:02:01 PDT

7SK RNA is a key player in the regulation of polymerase II transcription. 7SK RNA was considered as a highly conserved vertebrate innovation. The discovery of poorly conserved homologs in several insects and lophotrochozoans, however, implies a much earlier evolutionary origin. The mechanism of 7SK function requires interaction with the proteins HEXIM and LARP7. Here, we present a comprehensive computational analysis of these two proteins in metazoa, and we extend the collection of 7SK RNAs by several additional candidates. In particular we describe 7SK homologs in Caenorhabditis species. Furthermore, we derive an improved secondary structure model of 7SK RNA, which shows that the structure is quite well-conserved across animal phyla despite the extreme divergence at sequence level.

Tracing the history of goat pastoralism: new clues from mitochondrial and Y chromosome DNA in North Africa

Thu, 03 Sep 2009 05:03:31 PDT

Valuable insights into the history of human populations have been obtained by studying the genetic composition of their domesticated species. Here we address some of the long-standing questions about the origin and subsequent movements of goat pastoralism in Northern Africa. We present the first study combining results from mitochondrial DNA (mtDNA) and Y chromosome loci for the genetic characterization of a domestic goat population. Our analyses indicate a remarkably high diversity of maternal and paternal lineages in a sample of indigenous goats from the northwestern fringe of the African continent. Median-joining networks and a multidimensional scaling of ours and almost 2000 published mtDNA sequences revealed a considerable genetic affinity between goat populations from the Maghreb (Northwest Africa) and the Near East. It has been previously shown that goats have a weak phylogeographic structure compatible with high levels of gene flow, as demonstrated by the worldwide dispersal of the predominant mtDNA haplogroup A. In contrast, our results revealed a strong correlation between genetic and geographical distances in 20 populations from different regions of the world. The distribution of Y chromosome haplotypes in Maghrebi goats indicates a common origin for goat patrilines in both Mediterranean coastal regions. Taken together, these results suggest that the colonization and subsequent dispersal of domestic goats in Northern Africa was influenced by the maritime diffusion throughout the Mediterranean Sea and its coastal regions of pastoralist societies whose economy included goat herding. Finally, we also detected traces of gene flow between goat populations from the Maghreb and the Iberian Peninsula corroborating evidence of past cultural and commercial contacts across the Strait of Gibraltar.

Genic Incompatibilities in Two Hybrid Bacteriophages

Rokyta, D. R., Wichman, H. A. — Wed, 02 Sep 2009 06:45:52 PDT

Horizontal gene transfer and recombination play a major role in microbial evolution and have been detected in diverse groups, including many of medical relevance such as HIV and dengue virus. In the absence of mechanistic barriers, the evolutionary success of a particular recombination event is determined by whether the recombinant genotype suffers a fitness cost through the disruption of favorable epistatic interactions within the genome, and if so, the extent to which this fitness cost might be mitigated by subsequent compensatory evolution. To investigate the importance of epistatic interactions between genes and the evolutionary viability of a homologous recombination event between diverged ancestral genotypes, we constructed two recombinant microvirid bacteriophages by exchanging their alleles of the gene encoding the coat protein. The coding sequences for this gene differ by approximately 8% at the amino-acid level and were interchanged between two ancestral phages related to X174 and well adapted to their culture conditions. Because the recombinant phages showed drastically reduced fitnesses, we further explored their evolutionary viability by subjecting replicate lines of each of them to selection. We found that all four lineages achieved fitnesses commensurate with ancestral fitnesses in as few as 60 generations, and on average, the first substitution accounted for more than half of the total fitness recovery. Fitness recovery required three to five substitutions in each lineage, and overall eight of the nine essential phage genes were involved, suggesting extensive epistatic interactions throughout the genome. Interestingly, the proteins with the most extensive and apparent physical interactions with the exchanged protein in the viral capsid did not appear to have much of a role in fitness recovery. This result appears to be a consequence of the conservation of the amino-acid residues involved in the interactions. It suggests that strong epistatic interactions are less important than weaker, transient ones in producing genic incompatibilities, because they preclude variability in the interacting regions of the proteins.

Evolutionary Trajectories of Primate Genes Involved in HIV Pathogenesis

Wed, 02 Sep 2009 06:45:52 PDT

The current availability of five complete genomes of different primate species allows the analysis of genetic divergence over the last 40 million years of evolution. We hypothesized that the inter-species differences observed in susceptibility to HIV-1 would be influenced by the long-range selective pressures on host genes associated with HIV-1 pathogenesis. We established a list of human genes (n = 140) proposed to be involved in HIV-1 biology and pathogenesis, and a control set of 100 random genes. We retrieved the orthologous genes from the genome of humans and of four non-human primates (Pan troglodytes, Pongo pygmaeus abeli, Macaca mulatta and Callithrix jacchus) and analyzed the nucleotide substitution patterns of this dataset using codon-based maximum likelihood procedures. In addition, we evaluated whether the candidate genes have been targets of recent positive selection in humans by analyzing HapMap Phase 2 SNPs genotyped in a region centered on each candidate gene. A total of 1064 sequences were used for the analyses. Similar median K_A/K_S values were estimated for the set of genes involved in HIV-1 pathogenesis and for control genes; 0.19 and 0.15, respectively. However, genes of the innate immunity had median values of 0.37 (P-value = 0.0001, compared to control genes), and genes of intrinsic cellular defense had K_A/K_S values around or greater than 1.0 (P-value = 0.0002). Detailed assessment allowed the identification of residues under positive selection in 13 proteins: AKT1, APOBEC3G, APOBEC3H, CD4, DEFB1, GML, IL4, IL8RA, L-SIGN/CLEC4M, PTPRC/CD45, Tetherin/BST2, TLR7, and TRIM5. A number of those residues are relevant for HIV-1 biology. The set of 140 genes involved in HIV-1 pathogenesis did not show a significant enrichment in signals of recent positive selection in humans (intraspecies selection). However, we identified within or near these genes 24 polymorphisms showing strong signatures of recent positive selection. Interestingly, the DEFB1 gene presented signatures of both inter-species positive selection in primates and intra-species recent positive selection in humans. The systematic assessment of long-acting selective pressures on primate genomes is a useful tool to extend our understanding of genetic variation influencing contemporary susceptibility to HIV-1.

A new family of ferritin genes from Lupinus luteus - comparative analysis of plant ferritins, their gene structure, and evolution

Strozycki, P. M., Szymanski, M., Szczurek, A., Barciszewski, J., Figlerowicz, M. — Wed, 02 Sep 2009 06:45:51 PDT

Ferritins are one of the most important elements of cellular machinery involved in iron management. Despite extensive studies conducted during the last decade many factors regulating the expression of ferritin genes in plants remain unknown. To broaden our knowledge about the mechanisms controlling ferritin production in plant cells, we have identified and characterized a new family of ferritin genes (from yellow lupine). We have also inventoried all available plant ferritins and their genes and subjected them to a complex bioinformatic analysis. It showed that the conservative structure of ferritin genes was established much earlier than it was thought before. The first introns in ferritin genes appeared already in green algae. The number and location of introns have been finally established in mosses, over 400 million years ago and are strictly preserved in all plants from bryofites to dicots. Comparison of ferritin gene promoters revealed that the 14bp long iron dependent regulatory sequence (IDRS), identified earlier in Arabidopsis and maize, is characteristic for all higher plants. Moreover, we found that a highly conserved IDRS can be extended up to 22 bp (extIDRS). Phylogenetic analysis of plant ferritins showed that polypeptides of the eudicot clade can be divided into two sub-classes (eudicot-1 and eudicot-2). Interestingly, we found that genes encoding proteins classified as eudicot-1 and eudicot-2 are equipped with class-specific promoters. This suggests that eudicot ferritins are structurally and perhaps functionally diverse. Based on the above observations we were able to identify conservative elements other than extIDRS within plant ferritin gene promoters (ELEM1-6). We also found E-boxes and iron-responsive sequence elements FeRE1 and 2, characteristically distributed within ferritin promoters. Because most of the identified conserved sequences are located within or in close proximity of extIDRS, we named this fragment of the plant ferritin gene promoter the Regulatory Element Rich Region (RERR).

mtDNA Data Indicates a Single Origin for Dogs South of Yangtze River, less than 16,300 Years Ago, from Numerous Wolves

Tue, 01 Sep 2009 11:18:17 PDT

There is no generally accepted picture of where, when, and how the domestic dog originated. Previous studies of mitochondrial DNA (mtDNA) have failed to establish the time and precise place of origin because of lack of phylogenetic resolution in the so far studied control region (CR), and inadequate sampling. We therefore analysed entire mitochondrial genomes for 169 dogs to obtain maximal phylogenetic resolution, and the CR for 1,543 dogs across the Old World for a comprehensive picture of geographical diversity. Hereby, a detailed picture of the origins of the dog can for the first time be suggested. We obtained evidence that the dog has a single origin in time and space, and an estimation of the time of origin, number of founders and approximate region, which also gives potential clues about the human culture involved. The analyses showed that dogs universally share a common homogenous gene pool containing 10 major haplogroups. However, the full range of genetic diversity, all 10 haplogroups, was found only in south-eastern Asia south of Yangtze River, and diversity decreased following a gradient across Eurasia, through 7 haplogroups in Central China, and 5 in North China and Southwest Asia, down to only 4 haplogroups in Europe. The mean sequence distance to ancestral haplotypes indicates an origin 5,400-16,300 years ago from at least 51 female wolf founders. These results indicate that the domestic dog originated in southern China less than 16,300 years ago, from several hundred wolves. The place and time coincide approximately with the origin of rice agriculture, suggesting that the dogs may have originated among sedentary hunter-gatherers or early farmers, and the numerous founders indicate that wolf taming was an important culture trait.

Global microsatellite content distinguishes humans, primates, animals, and plants

Fri, 28 Aug 2009 05:29:52 PDT

Microsatellites are highly mutable, repetitive sequences commonly used as genetic markers, but they have never been studied en masse. Using a custom microarray to measure hybridization intensities of every possible repetitive nucleotide motif from 1-mers to 6-mers, we examined 25 genomes. Here we show that global microsatellite content varies predictably by species, as measured by array hybridization signal intensities, correlating with established taxonomic relationships, and particular motifs are characteristic of one species versus another. For instance, hominid-specific microsatellite motifs were identified despite alignment of the human reference, Celera, and Venter genomic sequences indicating substantial variation (30-50%) among individuals. Differential microsatellite motifs were mainly associated with genes involved in developmental processes, while those found in intergenic regions exhibited no discernible pattern. This is the first description of a method for evaluating microsatellite content to classify individual genomes.

A broad-scale phylogenetic analysis of group II intron RNAs and intron-encoded reverse transcriptases

Simon, D. M., Kelchner, S. A., Zimmerly, S. — Thu, 27 Aug 2009 06:54:41 PDT

Group II introns are self-splicing RNAs that are frequently assumed to be the ancestors of spliceosomal introns. They are widely distributed in bacteria and are also found in organelles of plants, fungi, and protists. In this study, we present a broad-scale phylogenetic analysis of group II introns using sequence data from both the conserved RNA structure and the intron-encoded reverse transcriptase (RT). Two similar phylogenies are estimated for the RT ORF, based on either amino acid or nucleotide sequence, while one phylogeny is produced for the RNA. In making these estimates, we confronted nearly all of the classic challenges to phylogenetic inference, including positional saturation, base composition heterogeneity, short internodes with low support, and sensitivity to taxon sampling. While the major lineages are well-defined, robust resolution of topology is not possible between these lineages. The approximately unbiased (AU) and Shimodaira-Hasegawa (SH) topology tests indicated that the RT ORF and RNA ribozyme data sets are in significant conflict under a variety of models, revealing the possibility of imperfect coevolution between group II introns and their intron-encoded ORFs. The high level of sequence divergence, large time scale, and limited number of alignable characters in our study are representative of many reverse transcriptases and group I introns, and our results suggest that phylogenetic analyses of any of these sequences could suffer from the same sources of error and instability identified in this study.

A phylogenomic approach to resolve the basal pterygote divergence

Simon, S., Strauss, S., von Haeseler, A., Hadrys, H. — Thu, 27 Aug 2009 06:54:41 PDT

One of the most fascinating Bauplan transitions in the animal kingdom was the invention of insect wings, a change which also contributed to the success and enormous diversity of this animal group. However, the origin of insect flight and the relationships of basal winged insect orders are still controversial. Three hypotheses have been proposed to explain the phylogeny of winged insects: (i) the traditional Palaeoptera hypothesis (Ephemeroptera+Odonata, Neoptera), (ii) the Metapterygota (Ephemeroptera, Odonata+Neoptera) and (iii) the Chiastomyaria hypothesis (Odonata, Ephemeroptera+Neoptera). Neither phylogenetic analyses of single genes nor even multiple marker systems (e.g. molecular markers + morphological characters) have yet been able to conclusively resolve basal pterygote divergences. A possible explanation for the lack of resolution is that the divergences took place in the mid-Devonian within a short period of time, and attempts to solve this problem have been confounded by the major challenge of finding molecular markers to accurately track these short ancient internodes. Although phylogenomic data are available for Neoptera and some wingless (apterygote) orders, they are lacking for the crucial Odonata and Ephemeroptera orders. We adopt a multi-gene approach including data from two new EST projects – from the orders Ephemeroptera (Baetis sp.) and Odonata (Ischnura elegans) – to evaluate the potential of phylogenomic analyses in clarifying this unresolved issue. We analyzed two data sets that differed in represented taxa, genes and overall sequence lengths: maxspe (15 taxa, 125 genes, 31,643 amino acid positions), maxgen (8 taxa, 150 genes, 42,541 amino acid positions). Maximum likelihood and Bayesian inference analyses both place the Odonata at the base of the winged insects. Furthermore, statistical hypotheses testing rejected both the Palaeoptera and the Metapterygota hypotheses. The comprehensive molecular data set developed here provides conclusive support for odonates as the most basal winged insect order (Chiastomyaria hypothesis). Data quality assessment indicates that proteins involved in cellular processes and signaling harbor the most informative phylogenetic signal.

Targets of balancing selection in the human genome

Thu, 27 Aug 2009 06:54:40 PDT

Balancing selection is potentially an important biological force for maintaining advantageous genetic diversity in populations, including variation that is responsible for long-term adaptation to the environment. By serving as a means to maintain genetic variation, it may be particularly relevant to maintaining phenotypic variation in natural populations. Nevertheless, its prevalence and specific targets in the human genome remain largely unknown. We have analyzed the patterns of diversity and divergence of 13,400 genes in two human populations using an unbiased single-nucleotide polymorphism (SNP) dataset, a genome-wide approach, and a method that incorporates demography in neutrality tests. We identified an unbiased catalog of genes with signatures of long-term balancing selection, which includes immunity genes as well as genes encoding keratins and membrane channels, and shows enrichment in functional categories involved in cellular structure. Patterns are mostly concordant in the two populations, with a small fraction of genes showing population-specific signatures of selection. Power considerations indicate that our findings represent a subset of all targets in the genome, suggesting that while balancing selection may not have an obvious impact on a large proportion of human genes, it is a key force affecting the evolution of a number of genes in humans.

Signal conflicts in the phylogeny of the primary photosynthetic eukaryotes

Deschamps, P., Moreira, D. — Tue, 25 Aug 2009 07:07:12 PDT

It is widely accepted that the first photosynthetic eukaryotes arose from a single primary endosymbiosis of a cyanobacterium in a phagotrophic eukaryotic host, which led to the emergence of three major lineages: Chloroplastida (green algae and land plants), Rhodophyta and Glaucophyta. For a long time, Glaucophyta have been thought to represent the earliest branch among them. However, recent massive phylogenomic analyses of nuclear genes have challenged this view, since most of them suggested a basal position of Rhodophyta, though with moderate statistical support. We have addressed this question by phylogenomic analysis of a large dataset of 124 proteins transferred from the chloroplast to the nuclear genome of the three Archaeplastida lineages. In contrast to previous analyses, we found strong support for the basal emergence of the Chloroplastida and the sister-group relationship of Glaucophyta and Rhodophyta. Moreover, the reanalysis of chloroplast gene sequences using methods more robust against compositional and evolutionary rate biases sustained the same result. Finally, we observed that the basal position of Rhodophyta found in the phylogenies based on nuclear genes depended on the sampling of sequences used as outgroup. When eukaryotes supposed to have never had plastids (animals and fungi) were used, the analysis strongly supported the early emergence of Glaucophyta instead of Rhodophyta. Therefore, there is a conflicting signal between genes of different evolutionary origins supporting either the basal branching of Glaucophyta or of Chloroplastida within the Archaeplastida. This second possibility would agree with the existence of the subkingdom Biliphyta, joining Glaucophyta and Rhodophyta.

Benchmarking next-generation transcriptome sequencing for functional and evolutionary genomics

Tue, 25 Aug 2009 07:07:11 PDT

Next-generation sequencing has opened the door to genomic analysis of non-model organisms. Technologies generating long sequence reads (200-400 bp) are increasingly used in evolutionary studies of non-model organisms, but the short sequence reads (30-50 bp) that can be produced at lower cost are thought to be of limited utility for de novo sequencing applications. Here, we tested this assumption by short-read sequencing the transcriptomes of the tropical disease vectors Aedes aegypti and Anopheles gambiae, for which complete genome sequences are available. Comparison of our results to the reference genomes allowed us to accurately evaluate the quantity, quality, and functional and evolutionary information content of our "test" data. We produced more than 0.7 billion nucleotides of sequenced data per species that assembled into more than 21,000 test contigs larger than 100 bp per species and covered ~27% of the Aedes reference transcriptome. Remarkably, the substitution error rate in the test contigs was ~0.25% per site, with very few indels or assembly errors. Test contigs of both species were enriched for genes involved in energy production and protein synthesis, and underrepresented in genes involved in transcription and differentiation. Ortholog prediction using the test contigs was accurate across hundreds of millions of years of evolution. Our results demonstrate the considerable utility of short-read transcriptome sequencing for genomic studies of non-model organisms, and suggest an approach for assessing the information content of next-generation data for evolutionary studies.

Transcription, Translation and the Evolution of Specialists and Generalists

Zhong, S., Miller, S. P., Dykhuizen, D. E., Dean, A. M. — Tue, 25 Aug 2009 07:07:11 PDT

We used DNA microarrays to measure transcription and iTRAQ 2D LC-MS/MS to measure protein expression in 14 strains of Escherichia coli adapted for hundreds of generations to growth-limiting concentrations of either lactulose, methylgalactoside or a 72:28 mixture of the two. The two ancestors, TD2 and TD10, differ only in their lac operons and have similar transcription and protein expression profiles. Changes in transcription and protein expression are observed at 30-250 genes depending on the evolved strain. Lactulose specialists carry duplications of the lac operon and show increased transcription and translation at lac. Methylgalactoside specialists are galS^– and so constitutively transcribe and translate mgl, which encodes a transporter of methylgalactoside. Two strains carry lac duplications, are galS^–, and show increased transcription and translation at both operons. One is a generalist, the other a lactulose specialist. The generalist fails to sweep to fixation because its lac⁺,galS⁺ competitor expresses the csg adhesin and sticks to the chemostat wall, thereby preventing complete wash-out. Transcription and translation are sometimes decoupled. Lactulose adapted strains show increased protein expression at fru, a fructose transporter, without evidence of increased transcription. This suggests that fructose, produced by the action of β-galactosidase on lactulose, may leach from cells before being recouped. Reduced expression, at ‘late’ flagella genes and the constitutive gat operon, is an adaptation to starvation. A comparison with two other long-term evolution experiments suggests that certain aspects of adaptation are predictable, some are characteristic of an experimental system, while others seem erratic.

Hemocyanin suggests a close relationship of Remipedia and Hexapoda

Ertas, B., von Reumont, B. M., Wagele, J.-W., Misof, B., Burmester, T. — Wed, 19 Aug 2009 11:35:19 PDT

The remipedia are enigmatic crustaceans from anchialine cave systems, first described only 30 years ago, whose phylogenetic affinities are as yet unresolved. Here we report the sequence of hemocyanin from Speleonectes tulumensis Yager, 1987 (Remipedia, Speleonectidae). This is the first proof of the presence of this type of respiratory protein in a crustacean taxon other than Malacostraca. S. tulumensis hemocyanin consists of multiple distinct (at least three) subunits (StuHc1 to 3). Surprisingly, the sequences are most similar to hexapod hemocyanins. Phylogenetic analyses showed that the S. tulumensis hemocyanin subunits StuHc1 and StuHc3 associate with the type 1 hexapod hemocyanin subunits, whereas StuHc2 associates with the type 2 subunits of hexapods. Together, remipede and hexapod hemocyanins are in the sister-group position to the hemocyanins of malacostracan crustaceans. Hemocyanins provide no indication of a close relationship of Myriapoda and Hexapoda, but tend to support Pancrustacea (Crustacea + Hexapoda). Our results also indicate that Crustacea are paraphyletic and that Hexapoda may have evolved from a Remipedia-like ancestor. Thus, Remipedia occupy a key position for the understanding of the evolution of hexapods, which are and have been one of the world's most speciose lineage of animals.

Phylogeny of the "forgotten" cellular slime mold, Fonticula alba, reveals a key evolutionary branch within Opisthokonta

Brown, M. W., Spiegel, F. W., Silberman, J. D. — Wed, 19 Aug 2009 11:35:18 PDT

The shared ancestry between Fungi and animals has been unequivocally demonstrated by abundant molecular and morphological data for well over a decade. Along with the animals and Fungi, multiple protists have been placed in the supergroup Opisthokonta making it exceptionally diverse. In an effort to place the cellular slime mold Fonticula alba, an amoeboid protist with aggregative, multicellular fruiting, we sequenced five nuclear encoded genes; small subunit ribosomal RNA, actin, beta-tubulin, elongation factor 1-alpha, and the cytosolic isoform of heat shock protein 70 for phylogenetic analyses. Molecular trees demonstrate that Fonticula is an opisthokont that branches sister to filose amoebae in the genus Nuclearia. Fonticula plus Nuclearia are sister to Fungi. We propose a new name for this well-supported clade, Nucletmycea, incorporating Nuclearia, Fonticula, and Fungi. Fonticula represents the first example of a cellular slime mold morphology within Opisthokonta. Thus, there are four types of multicellularity in the supergroup - animal, fungal, colonial, and now aggregative. Our data indicate that multicellularity in Fonticula evolved independent of that found in the fungal and animal radiations. With the rapidly expanding sequence and genomic data becoming available from many opisthokont lineages, Fonticula may be fundamental to understanding opisthokont evolution as well as any possible commonalities involved with the evolution of multicellularity.

General Heterotachy and Distance Method Adjustments

Wu, J., Susko, E. — Mon, 17 Aug 2009 12:07:47 PDT

Heterotachy is a general term to describe positions in a sequence that evolve at different rates in different lineages. Kolaczkowski and Thornton (2004) recently described an intriguing heterotachy model that leads to topological bias for likelihood-based methods and parsimony methods. In this article, we show that heterotachy can generally be viewed as multivariate rates-across-sites variation, which can be described as randomly drawing rates (or branch lengths) from a multivariate distribution for each branch at each site. Motivated by this idea, we propose a pairwise alpha heterotachy adjustment model, which gives us much improved topological estimation in the Kolaczkowski and Thornton (2004) settings.

Molecular evolution of GYPC: Evidence for recent structural innovation and positive selection in humans

Wilder, J. A., Hewett, E. K., Gansner, M. E. — Thu, 13 Aug 2009 11:14:11 PDT

GYPC encodes two erythrocyte surface sialoglycoproteins in humans, glycophorin C and glycophorin D (GPC & GPD), via initiation of translation at two start codons on a single transcript. The malaria-causing parasite Plasmodium falciparum uses GPC as a means of invasion into the human red blood cell. Here we examine the molecular evolution of GYPC among the Hominoidea (Greater and Lesser Apes) and also the pattern of polymorphism at the locus in a global human sample. We find an excess of non-synonymous divergence among species that appears to be caused solely by accelerated evolution of GYPC in the human lineage. Moreover, we find that the ability of GYPC to encode both GPC and GPD is a uniquely human trait, caused by the evolution of the GPC start codon in the human lineage. The pattern of polymorphism among humans is consistent with a hitchhiking event at the locus, suggesting that positive natural selection affected GYPC in the relatively recent past. Because GPC is exploited by P. falciparum for invasion of the red blood cell, we hypothesize that selection for evasion of P. falciparum has caused accelerated evolution of GYPC in humans (relative to other primates), and that this positive selection has continued to act in the recent evolution of our species. These data suggest that malaria has played a powerful role in shaping molecules on the surface of the human red blood cell. In addition, our examination of GYPC reveals a novel mechanism of protein evolution: co-option of UTR sequence following the formation of a new start codon. In the case of human GYPC the ancestral protein (GPD) continues to be produced through leaky translation. Because leaky translation is a widespread phenomenon among genes and organisms, we suggest that co-option of UTR sequence may be an important source of protein innovation.