Molecular Biology and Evolution - Advance Access

Genealogical discontinuities among Etruscan, Medieval and contemporary Tuscans

2009-07-01

The available mitochondrial DNA (mtDNA) data do not point to clear genetic relationships between current Tuscans and the Bronze-Age inhabitants of Tuscany, the Etruscans. To understand how and when such a genetic discontinuity may have arisen, we extracted and typed the mtDNAs of 27 medieval Tuscans from an initial sample of 61, spanning a period between the 10^th and 15^th centuries A.D.. We then tested by serial coalescent simulation various models describing the genealogical relationships among past and current inhabitants of Tuscany, the latter including three samples (from Murlo, Volterra, Casentino) which were recently claimed to be of Etruscan descent. Etruscans and medieval Tuscans share three mitochondrial haplotypes, but fall in distinct branches of the mitochondrial genealogy in the only model that proved compatible with the data. Under that model, contemporary people of Tuscany show clear genetic relationships with Medieval people, but not with the Etruscans, along the female lines. No evidence of excess mutation was found in the Etruscan DNAs by a Bayesian test, and so there is no reason to suspect that these results be biased by systematic contamination of the ancient sequences or laboratory artefacts. Extensive demographic changes before 1000 A.D. are thus the simplest explanation for the differences between the contemporary and the Bronze-Age mitochondrial DNAs of Tuscany. Accordingly, genealogical continuity between ancient and modern populations of the same area does not seem a safe general assumption, but rather a hypothesis that, when possible, should be tested using ancient DNA analysis.

The consequences of base pair composition biases for regulatory network organization in prokaryotes

Cordero, O. X., Hogeweg, P. — 2009-06-30

Given the dramatic variation GC content observed in prokaryotes, from ~ 20% to ~ 75% GC, one wonders if these extreme biases in base pair composition affect the evolution of transcription factor binding sites. This letter shows that, along the wide range of GC content variation in bacteria, bacterial binding sites keep a high frequency of AT bases, roughly independently of the background base pair composition of intergenic regions. As a result, the equilibrium base pair frequencies of binding sites depart the most from those of background DNA in GC rich genomes. This not only implies a higher specificity, but also a higher coding barrier for binding sites in GC rich genomes. In accordance, we observe that the average percentage of divergently transcribed regions increases with the GC content of the genome, suggesting the use of a more efficient coding strategy.

Application of the isolation with migration model demonstrates the Pleistocene origin of geographic differentiation in Cardamine nipponica (Brassicaceae), an endemic Japanese alpine plant

Ikeda, H., Fujii, N., Setoguchi, H. — 2009-06-30

The Pleistocene was characterized by a cyclic pattern of cold and warm climatic periods, or climatic oscillations, which caused fluctuations in the distributions of organisms. This resulted in drastic changes in demography, thereby accelerating the genetic divergence of populations. Phylogeographic studies have elucidated the history of populations during the Pleistocene. However, given the lack of model-based analysis of population histories, previous phylogeographic studies could not adequately evaluate the effect of these Pleistocene climatic oscillations on the genetic divergence and migration events between populations. Populations of Japanese alpine plants in central and northern Japan are highly differentiated, and a history of isolation between regions during the Pleistocene was inferred. Using sequences of ten nuclear genes (approximately 7,000 bp in total) from Cardamine nipponica (Brassicaceae), we applied an isolation with migration (IM) model to test the significance of the isolation history between central and northern Japan and to assess whether range shifts during the Pleistocene climatic oscillations were involved in the genetic differentiation between regions. The estimated divergence time indicates that the two regions were separated about 100,000–110,000 years ago. The exclusive occurrence of closely related haplotypes within each region (parsimony network) and the high level of genetic differentiation between the regions (mean F_ST = 0.417) indicate that genetic divergence occurred following the isolation of the two regions. Therefore, the genetic differentiation between regions was shaped during the Pleistocene, especially during the last glacial and inter- and post-glacial periods. In addition, our multilocus analysis showed that populations in central and northern Japan were completely isolated after they split. Geographic separation and subsequent restricted migration events among mountains could explain this isolation history between regions. Furthermore, genetic drift in the reduced populations would remove evidence of occasional migration, emphasizing the isolation history. Therefore, our application of a demographic model demonstrated the Pleistocene origin of geographic differentiation statistically and provided a plausible migration history for C. nipponica.

Streamlining and large ancestral genomes in Archaea inferred with a phylogenetic birth-and-death model

Csuros, M., Miklos, I. — 2009-06-30

Homologous genes originate from a common ancestor through vertical inheritance, duplication or horizontal gene transfer. Entire homolog families spawned by a single ancestral gene can be identified across multiple genomes based on protein sequence similarity. The sequences, however, do not always reveal conclusively the history of large families. In order to study the evolution of complete gene repertoires, we propose here a mathematical framework that does not rely on resolved gene family histories. We show that so-called phylogenetic profiles, formed by family sizes across multiple genomes, are sufficient to infer principal evolutionary trends. The main novelty in our approach is an efficient algorithm to compute the likelihood of a phylogenetic profile in a model of birth-and-death processes acting on a phylogeny.

We examine known gene families in 28 archaeal genomes using a probabilistic model that involves lineage- and family-specific components of gene acquisition, duplication, and loss. The model enables us to consider all possible histories when inferring statistics about archaeal evolution. According to our reconstruction, most lineages are characterized by a net loss of gene families. Major increases in gene repertoire have occurred only a few times. Our reconstruction underlines the importance of persistent streamlining processes in shaping genome composition in Archaea. It also suggests that early archaeal genomes were as complex as typical modern ones, and even show signs, in the case of the methanogenic ancestor, of an extremely large gene repertoire.

Haplotype Sharing Analysis Showing Uyghurs Are Unlikely Genetic Donors

Xu, S., Jin, W., Jin, L. — 2009-06-29

The Uyghur are a group of people primarily residing in Xinjiang of China which is geographically located in Central Asia, from where modern humans were presumably spread in all directions reaching Europe, east and northeast Asia about 40 kya. A recent study suggested that the Uyghur are ancestry donors of the East Asian gene pool. However, an alternative hypothesis, i.e. the Uyghur is an admixture population with both East Asian (EAS) and European (EUR) ancestries is also supported by our previous studies. To test the two competing hypotheses, here we conducted a haplotype sharing analysis based on empirical and simulated data of high density single nucleotide polymorphisms (SNPs). Our results showed that more than 95% of Uyghur (UIG) haplotypes could be found in either East Asian (EAS) or European (EUR) populations, which contradicts the expectation of the null models assuming that UIG are donors. Simulation studies further indicated that the proportion of UIG private haplotypes observed in empirical data is only expected in alternative models assuming that UIG is an admixture population. Interestingly, the estimated ancestry contribution of 44%:56% (EAS:EUR) based on haplotype sharing analysis is consistent with our previous estimation with STRUCTURE analysis. Although the history of Uyghurs could be complex, our method is explicit and conservative in rejecting the null hypothesis. We concluded that the gene pool of modern Uyghurs is more likely a sole recipient with contribution from both EAS and EUR.

CGIN1: a retroviral contribution to mammalian genomes

Marco, A., Marin, I. — 2009-06-26

This study describes the origin and structural features of a mammalian gene, CGIN1 (Cousin of GIN1). CGIN1 proteins contain a NYN domain, retroviral RNase H and integrase domains and a domain of unknown function (CGIN1 domain) which is also present in two other genes (N4BP1 and KIAA0323). We suggest that CGIN1 derives from the fusion of a KIAA0323-like gene with retroviral sequences, which occurred prior to the marsupial/eutherian split. Sequence and structural analyses indicate that the CGIN1 integrase domain is inactive, but still retains the three-dimensional folding observed in retroviral integrases. We hypothesize that CGIN1 may contribute to retroviral resistance in mammals by regulating the ubiquitination of viral proteins.

Lack of Character Displacement in the Male Recognition Molecule, Bindin, in Altantic Sea Urchins of the Genus Echinometra

Geyer, L. B., Lessios, H. A. — 2009-06-26

Bindin, a protein involved in sea urchin sperm-egg recognition and adhesion, is under positive selection in genera with sympatric species but evolves neutrally in genera in which all species are allopatric. This pattern has led to suggestions that reinforcement may be the source of the observed selection. Reproductive character displacement, or increased divergence of reproductive characters in areas where closely related species overlap, is often a consequence of reinforcement and has been shown to be present in one Indo-Pacific species of the genus Echinometra. In the Atlantic species of the same genus, positive selection has been shown to act on bindin of Echinometra lucunter. To examine whether the source of this selection is reinforcement, we determined variation on the first exon of bindin in E. lucunter in the Caribbean, where it is sympatric with E. viridis, and in the rest of the Atlantic, where E. viridis is absent. There was no differentiation between bindin sequences from the two geographic regions; similar levels of positive selection were found to be acting in both areas. The similarities were not due to gene flow; mtDNA from the two regions indicates that E. lucunter populations most likely originated in the Atlantic and have not exchanged genes with Caribbean populations for approximately 200,000 years. The lack of evidence of stronger selection on bindin of E. lucunter in areas of sympatry with its sister species suggests that the source of selection is not reinforcement. Processes acting within species, such as sexual selection, sperm competition or sexual conflict, are more likely to be involved in the evolution of this molecule.

Comparative evolutionary analysis of the major structural subunit of Vibrio vulnificus type IV pili

2009-06-25

Type IV pili contribute to virulence in Vibrio vulnificus, the bacterium responsible for the majority of fatal seafood-related infections. Here we performed within- and between-species evolutionary analysis of the gene that encodes the major structural subunit of the pilus, pilA, by comparing it with pilD and gyrB, the genes encoding the type IV prepilin peptidase and β subunit of DNA gyrase respectively. While the diversity in pilD and gyrB is similar to each other and likely to have accumulated after speciation of V. vulnificus, pilA is several times more diverse at both nonsynonymous and synonymous levels. Also, in contrast to pilD and gyrB, there are virtually unrestricted and highly-localized horizontal movements of pilA alleles between the major phylogenetic groups of V. vulnificus. The frequent movement of pilA involves homologous recombination of the entire gene with no evidence for intragenic recombination between the alleles. We propose that pilA allelic diversity and horizontal movement is maintained in the population by both diversifying and frequency-dependent selection most likely to escape shellfish innate immunity defense or lytic phages. Other possibilities leading to such selection dynamics of V. vulnificus pilA could involve adaptation to diverse host populations or within-host compartments, or natural DNA uptake and transformation. We show that the history of nucleotide diversification in pilA predates V. vulnificus speciation and this diversification started at or before the time of the last common ancestor for V. vulnificus, V. parahaemolyticus and V. cholerae. At the same time, it appears that within the various pilA groups of V. vulnificus there is no positive selection for structural mutations and consequently no evidence for source-sink selection. In contrast, pilD has accumulated a number of apparently adaptive mutations in the regions encoding the membrane-spanning portions of the prepilin peptidase, possibly affecting fimbrial expression and/or function, and is being subjected to source-sink selection dynamics.

Evolutionary origin and functions of retrogene introns

Fablet, M., Bueno, M., Potrzebowski, L., Kaessmann, H. — 2009-06-24

Retroposed genes (retrogenes) originate via the reverse transcription of mature messenger RNAs from parental source genes and are therefore usually devoid of introns. Here we characterize a particular set of mammalian retrogenes that acquired introns upon their emergence and thus represent rare cases of intron gain in mammals. We find that while a few retrogenes evolved introns in their coding or 3’ untranslated regions, most introns originated together with untranslated (UTR) exons in the 5’ flanking regions of the retrogene insertion site. They emerged either de novo or through fusions with 5’ UTR exons of host genes into which the retrogenes inserted. Generally, retrogenes with introns display high transcription levels and show broader spatial expression patterns than other retrogenes. Our experimental expression analyses of individual intron–containing retrogenes show that 5’ UTR introns may indeed promote higher expression levels, at least in part through encoded regulatory elements. By contrast, 3’ UTR introns may lead to down–regulation of expression levels via nonsense–mediated decay mechanisms. Notably, the majority of retrogenes with introns in their 5’ flanks depend on distant, sometimes bidirectional CpG dinucleotide–enriched promoters for their expression that may be recruited from other genes in the genomic vicinity. We thus propose a scenario where the acquisition of new 5’ exon–intron structures was directly linked to the recruitment of distant promoters by these retrogenes, a process potentially facilitated by the presence of proto–splice sites in the genomic vicinity of retrogene insertion sites. Thus, the primary role and selective benefit of new 5’ introns (and UTR exons) was probably initially to span the often substantial distances to potent CpG promoters driving retrogene transcription. Later in evolution, these introns then obtained additional regulatory roles in fine–tuning retrogene expression levels. Our study provides novel insights regarding mechanisms underlying the origin of new introns, the evolutionary relevance of intron gain, and the origin of new gene promoters.

Protease Gene Duplication and Proteolytic Activity in Drosophila Female Reproductive Tracts

Kelleher, E. S., Pennington, J. E. — 2009-06-22

Secreted proteases play integral roles in sexual reproduction in a broad range of taxa. In the genetic model Drosophila melanogaster, these molecules are thought to process peptides and activate enzymes inside female reproductive tracts, mediating critical post-mating responses. A recent study of female reproductive tract proteins in the cactophilic fruit-fly D. arizonae, identified pervasive, lineage-specific gene duplication amongst secreted proteases. Here we compare the evolutionary dynamics, biochemical nature, and physiological significance of secreted female reproductive serine endoproteases (SFRSEs) between D. arizonae and its congener D. melanogaster. We show that D. arizonae lower female reproductive tract (LFRT) proteins are significantly enriched for recently-duplicated secreted proteases, particularly serine endoproteases, relative to D. melanogaster. Isolated lumen from D. arizonae LFRTs, furthermore, exhibits significant tryspin-like and elastase-like serine endoprotease acitivity, while no such activity is seen in D. melanogaster. Finally, trypsin and elastase-like activity in D. arizonae female reproductive tracts is negatively regulated by mating. We propose that the intense proteolytic environment of the D. arizonae female reproductive tract relates to the extraordinary reproductive physiology of this species, and that ongoing gene duplication amongst these proteases is an evolutionary consequence of sexual conflict.

Large gene family expansions and adaptive evolution for odorant and gustatory receptors in the pea aphid, Acyrthosiphon pisum

Smadja, C., Shi, P., Butlin, R. K., Robertson, H. M. — 2009-06-19

Gaining insight into the mechanisms of chemoreception in aphids is of primary importance for both integrative studies on the evolution of host plant specialization and applied research in pest control management since aphids rely on their sense of smell and taste to locate and assess their host plants. We made use of the recent genome sequence of the pea aphid, Acyrthosiphon pisum, to address the molecular characterization and evolution of key molecular components of chemoreception: the odorant (Or) and gustatory (Gr) receptor genes. We identified 79 Or and 77 Gr genes in the pea aphid genome and showed that most of them are aphid-specific genes that have undergone recent and rapid expansion in the genome. By addressing selection within sets of paralogous Or and Gr expansions, for the first time in an insect species, we show that the most recently duplicated loci have evolved under positive selection, which might be related to the high degree of ecological specialization of this species. While more functional studies are still needed for insect chemoreceptors, we provide evidence that Grs and Ors have different sets of positively selected sites, suggesting the possibility that these two gene families might have different binding pockets and bind structurally distinct classes of ligand. The pea aphid is the most basal insect species with a completely sequenced genome to date. The identification of chemoreceptor genes in this species is a key step towards further exploring insect comparative genetics, the genomics of ecological specialization and speciation, and new pest control strategies.

Comparison of queen-specific gene expression in related lower termite species

Weil, T., Korb, J., Rehli, M. — 2009-06-18

The molecular mechanisms regulating caste determination and reproductive division of labor, the hallmarks of insect societies, are poorly defined. The identification of key genes involved in these developmentally important processes will be essential to gain a better understanding of the mechanisms controlling one of the most impressive examples of polyphenism, the caste structure of eusocial species. Here we applied representational difference analysis (RDA) of cDNAs, to study differential gene expression between queens (female neotenics) and workers in the drywood termite Cryptotermes cynocephalus and identified thirteen genes that were highly expressed in queens. In addition, we partially cloned several homologous genes of the related termite species Cryptotermes secundus, and compared the expression profiles of ten homologous genes. In most cases the preferential expression in female neotenics was not conserved between species, despite the close phylogenetic relationship of both Cryptotermes species. It is possible that these genes are associated with known species-specific differences in caste development modes. Only three genes (Neofem1, 2, and 3) showed a conserved and highly preferential expression in female neotenics, suggesting that their products may play important roles in female reproductives, in particular in controlling caste determination and reproductive division of labor.

On the origins and admixture of Malagasy: new evidence from high resolution analyses of paternal and maternal lineages

2009-06-17

The Malagasy have been shown to be a genetically admixed population combining parental lineages with African and South East Asian ancestry. In the present paper we fit the Malagasy admixture history in a highly resolved phylogeographic framework by typing a large set of mtDNA and Y DNA markers in unrelated individuals from inland (Merina) and coastal (Antandroy, Antanosy, Antaisaka) ethnic groups. This allowed performance of a multi-level analysis in which the diversity among main ethnic divisions, lineage ancestries, and modes of inheritance could be concurrently evaluated. Admixture was confirmed to result from the encounter of African and South-East Asian people with minor recent male contributions from Europe. However, new scenarios are depicted about Malagasy admixture history. The distribution of ancestral components was ethnic- and sex-biased, with the Asian ancestry appearing more conserved in the female than in the male gene pool, and in inland than in coastal groups. A statistic based on haplotype sharing (D_HS), showing low sampling error and time linearity over the last 200 generations, was here introduced for the first time and helped to integrate our results with linguistic and archeological data. The focus about the origin of Malagasy lineages was enlarged in space and pushed back in time. Homelands could not be pinpointed, but appeared to comprise two vast areas containing different populations from sub-Saharan Africa and South-East Asia. The pattern of diffusion of uni-parental lineages was compatible with at least two events: a primary admixture of proto-Malay people with Bantu-speakers bearing a western-like pool of haplotypes, followed by a secondary flow of Southeastern Bantu-speakers unpaired for gender (mainly male-driven) and geography (mainly coastal).

Estimating the rate of adaptive molecular evolution in the presence of slightly deleterious mutations and population size change

Eyre-Walker, A., Keightley, P. D. — 2009-06-17

The prevalence of adaptive evolution relative to genetic drift is a central problem in molecular evolution. Methods to estimate the fraction of adaptive nucleotide substitutions () have been developed, based on the McDonald-Kreitman test, that contrast polymorphism and divergence between selectively and neutrally evolving sites. However, these methods are expected to give downwardly biased estimates of if there are slightly deleterious mutations, because these inflate polymorphism relative to divergence. Here, we estimate by simultaneously estimating the distribution of fitness effects of new mutations at selected sites from the site frequency spectrum, and the number of adaptive substitutions. We test the method using simulations. If data meet the assumptions of the analysis model, estimates of show little bias, even when there is little or no recombination. However, population size differences between the divergence and polymorphism phases may cause to be over- or under-estimated by a predictable factor that depends on the magnitude of the population size change and the shape of the distribution of effects of deleterious mutations. We analyse several data sets of protein-coding genes and noncoding regions from hominids and Drosophila. In Drosophila genes, we estimate that c.50% of amino acid substitutions and c.20% of substitutions in introns are adaptive. In protein-coding and noncoding data sets of humans, comparison to macaque sequences reveals little evidence for adaptive substitutions. However, the true frequency of adaptive substitutions in human coding DNA could be as high as 40%, since estimates based on current polymorphism may be strongly downwardly biased by a decrease in the effective population size along the human lineage.

INTEGRATING Y-CHROMOSOME, MITOCHONDRIAL AND AUTOSOMAL DATA TO ANALYSE THE ORIGIN OF PIG BREEDS

2009-06-17

We have investigated the origin of swine breeds through the joint analysis of mitochondrial, microsatellite and Y-chromosome polymorphisms in a sample of pigs and wild boars with a worldwide distribution. Genetic differentiation between pigs and wild boars was remarkably weak, likely as a consequence of a sustained gene flow between both populations. The analysis of nuclear markers evidenced the existence of a close genetic relationship between Near Eastern and European wild boars making difficult to infer their relative contributions to the gene pool of modern European breeds. Moreover, we have shown that European and Far Eastern pig populations have contributed maternal and paternal lineages to the foundation of African and South American breeds. While West African pigs from Nigeria and Benin exclusively harboured European alleles, Far Eastern and European genetic signatures of similar intensity were detected in swine breeds from Eastern Africa. This region seems to have been a major point of entry of livestock species in the African continent as a result of the Indian Ocean trade. Finally, South American creole breeds had essentially a European ancestry although Asian Y-chromosome and mitochondrial haplotypes were found in a few Nicaraguan pigs. The existence of Spanish and Portuguese commercial routes linking Asia with America might have favoured the introduction of Far Eastern breeds into this continent.

The protein import channel in the outer mitosomal membrane of Giardia intestinalis

2009-06-16

The identification of mitosomes in Giardia generated significant debate on the evolutionary origin of these organelles; whether they were highly reduced mitochondria, or the product of a unique endosymbiotic event in an amitochondrial organism. As the protein import pathway is a defining characteristic of mitochondria, we sought to discover a TOM (translocase in the outer mitochondrial membrane) complex in Giardia. A Hidden Markov model search of the Giardia genome identified a Tom40 homologous sequence (GiTom40), where Tom40 is the protein translocation channel of the TOM complex. The GiTom40 protein is located in the membrane of mitosomes in a ~200 kDa TOM complex. As Tom40 was derived in the development of mitochondria to serve as the protein import channel in the outer membrane, its presence in Giardia evidences the mitochondrial ancestry of mitosomes.

The CKK domain (DUF1781) domain binds microtubules and defines the CAMSAP/ssp4 family of animal proteins

2009-06-09

We describe a structural domain common to proteins related to human CAMSAP1 (calmodulin-regulated spectrin-associated protein1). Analysis of the sequence of CAMSAP1 identified a domain near the C-terminus common to CAMSAP1 and two other mammalian proteins KIAA1078 and KIAA1053, which we term a CKK domain. This domain was also present in invertebrate CAMSAP1 homologues, and was found in all available eumetazoan genomes (including cnidaria), but not in the placozoan Trichoplax adherens, nor in any non-metazoan organism. Analysis of codon alignments by the sitewise likelihood ratio method gave evidence for strong purifying selection on all codons of mammalian CKK domains, potentially indicating conserved function. Interestingly, the Drosophila homologue of the CAMSAP family is encoded by the ssp4 gene, which is required for normal formation of mitotic spindles. To investigate function of the CKK domain, human CAMSAP1-EGFP and fragments including the CKK domain were expressed in HeLa cells. Both whole CAMSAP1 and the CKK domain showed localization coincident with microtubules. In vitro, both whole CAMSAP1-GST and CKK-GST bound to microtubules. Immunofluorescence using anti-CAMSAP1 antibodies on cerebellar granule neurons revealed a microtubule pattern. Over-expression of the CKK domain in PC12 cells blocked production of neurites, a process that requires microtubule function. We conclude that the CKK domain binds microtubules and represents a domain that evolved with the metazoa.

Positively selected disease response orthologous gene sets in the cereals identified using Sorghum bicolor L. Moench expression profiles and comparative genomics

Zamora, A., Sun, Q., Hamblin, M. T., Aquadro, C. F., Kresovich, S. — 2009-06-08

Disease response genes diverge under recurrent positive selection as a result of a molecular arms-race between hosts and pathogens. Most of these studies were conducted in animals, and few defense genes have been shown to evolve adaptively in plants. To test for adaptation in the molecules mediating disease resistance in the cereals we first combined information from the expression pattern of Sorghum bicolor genes and from divergence to the full genome of rice to identify candidate disease response genes. We then used evolutionary analyses of orthologous gene sets from several grass species, to determine whether the disease response genes show signals of positive selection and the residues targeted. We found 140 divergent genes upregulated under biotic stress in S. bicolor by evaluating the relative abundance of ESTs in different libraries and comparing them to rice genes. For 10 of these genes, we found sets of orthologs including sequences from rice and three other cereals; 6 genes showed a pattern of substitution that was consistent with positive selection. Three of these genes, a thaumatin, a peroxidase and a barley mlo homolog, are known antifungal proteins. The other three genes with evidence of positive selection were a MADS box transcription factor, an eIF5 translation initiation factor and a gene of unknown function but with evidence of expression during stress. Permutation analyses, using different ortholog and paralog sequences, consistently identified 5 positively selected codons in the peroxidase, a member of a cluster of genes and a large gene family. We mapped the positively selected residues onto the structure of the peroxidase and thaumatin and found that all sites are on the surface of these proteins and several are close to biochemically determined active sites. Identifying new positively selected plant disease resistance genes and the critical amino acid sites provides a basis for functional studies that may increase our understanding of their underlying molecular mechanisms of action. Additionally, it may lead to the identification of individuals having variation at functionally important sites, as well as eventually using this information in the rational design and engineering of proteins involved in plant disease resistance.

Genomic features that predict allelic imbalance in humans suggest patterns of constraint on gene expression variation

Tung, J., Fedrigo, O., Haygood, R., Mukherjee, S., Wray, G. A. — 2009-06-08

Variation in gene expression is an important contributor to phenotypic diversity within and between species. Although this variation often has a genetic component, identification of the genetic variants driving this relationship remains challenging. In particular, measurements of gene expression usually do not reveal whether the genetic basis for any observed variation lies in cis or in trans to the gene, a distinction that has direct relevance to the physical location of the underlying genetic variant, and which may also impact its evolutionary trajectory. Allelic imbalance measurements identify cis-acting genetic effects by assaying the relative contribution of the two alleles of a cis-regulatory region to gene expression within individuals. Identification of patterns that predict commonly imbalanced genes could therefore serve as a useful tool and also shed light on the evolution of cis-regulatory variation itself. Here, we show that sequence motifs, polymorphism levels, and divergence levels around a gene can be used to predict commonly imbalanced genes in a human dataset. Reduction of this feature set to four factors revealed that only one factor significantly differentiated between commonly imbalanced and non-imbalanced genes. We demonstrate that these results are consistent between the original dataset and a second published dataset in humans obtained using different technical and statistical methods. Finally, we show that variation in the single allelic imbalance-associated factor is partially explained by the density of genes in the region of a target gene (allelic imbalance is less probable for genes in gene-dense regions), and, to a lesser extent, the evenness of expression of the gene across tissues and the magnitude of negative selection on putative regulatory regions of the gene. These results suggest that the genomic distribution of functional cis-regulatory variants in the human genome is nonrandom, perhaps due to local differences in evolutionary constraint.

Viral resistance evolution fully escapes a rationally-designed lethal inhibitor

Keller, T. E., Molineux, I. J., Bull, J. J. — 2009-06-03

Viruses are notoriously capable of evolving resistance to drugs. However, if the endpoint of resistance evolution is only partial escape, a feasible strategy should to be stack drugs, so the combined effect of partial inhibition by several drugs results in net inhibition. Assessing the feasibility of this approach requires quantitative data on viral fitness before and after evolution of resistance to a drug, as done here with bacteriophage T7. An inhibitory gene expressed from a phage promoter aborts wild-type T7 infections. The effect is so severe that the phage population declines when exposed to the inhibitor but expands a billion-fold per hour in its absence. In prior work, T7 evolved modest resistance to this inhibitor, an expected result. Given the nature of the inhibitor, that it used the phage's own promoter to target the phage's destruction, we anticipated that resistance evolution would be limited as the phage may need to evolve a new regulatory system, with simultaneous changes in its RNA polymerase and many of its promoters to fully escape inhibition. We show here that further adaptation of the partially resistant phage led to complete resistance. Resistance evolution was due to three mutations in the RNA polymerase gene and two other genes; unexpectedly, no changes were observed in promoters. Consideration of other mechanisms of T7 inhibition leaves hope that permanent inhibition of viral growth with drugs can in principle be achieved.

Evolution of Vault RNAs

2009-06-02

Vault RNAs (vtRNAs) are small, about 100nt long, polymerase III transcripts contained in the vault particles of eukaryotic cells. Presumably due to their enigmatic function, they have received little attention compared to most other non-coding RNA (ncRNA) families. Their poor sequence conservation makes homology search a complex and tedious task even within vertebrates. Here we report on a systematic and comprehensive analysis of this rapidly evolving class of ncRNAs in deuterostomes, providing a comprehensive collection of computationally predicted vtRNA genes. We find that all previously described vtRNAs are located at a conserved genomic locus linked to the protocadherin gene cluster, an association that is conserved throughout gnathostomes. Lineage specific expansions to small vtRNA gene clusters are frequently observed in this region. A second vtRNA locus is syntenically conserved across eutherian mammals. The vtRNAs at the two eutherian loci exhibit substantial differences in their promoter structures, explaining their differential expression patterns in several human cancer cell lines. In teleosts, expression of several paralogous vtRNA genes, most but not all located at the syntenically conserved protocadherin locus, was verified by RT-PCR.

Low exchangeability of selenocysteine, the 21st amino acid, in vertebrate proteins

Castellano, S., Andres, A. M., Bosch, E., Bayes, M., Guigo, R., Clark, A. G. — 2009-06-01

Selenocysteine (Sec), the 21^st amino acid, is incorporated into proteins through the recoding of a termination codon, an inefficient translational process mediated by a complex molecular machinery. Sec is a rare amino acid in extant proteins, chemically similar to cysteine (Cys), found in homologous position to Cys of non-selenoprotein families. Selenoproteins account for the dependence of vertebrates on environmental selenium (Se) and have an important role in several Se-deficiency diseases. Selenoproteins are poorly characterized enzymes and reports on the functional exchangeability of Sec with Cys are limited and controversial. Whether the unique role of Sec in some selenoenzymes illustrates the broader contribution of selenium to protein function is unknown (Gromer et al. 2003, Proc Natl Acad Sci USA 100:12618; 10.1073/pnas.2134510100). Here, we address this question from an evolutionary perspective by the simultaneous identification of the patterns of divergence in almost half a billion years of vertebrate evolution and diversity within the human lineage for the full complement of enzymatic Sec residues in these proteomes. We complete this analysis with data for the homologous Cys residues in the same genomes. Our results indicate concerted purifying selection across Sec and Cys sites in all selenoproteomes, consistent with a unique role of Sec in protein function, low exchangeability and an unknown degree of functional divergence with Cys-homologs. The distinct biochemical properties of Sec, rather than the geographical distribution of selenium, global O₂ levels or Sec metabolic cost, appear to play a major role in driving adaptive changes in vertebrate selenoproteomes. A better understanding of the selenoproteomes and neutral evolutionary patterns in other taxa will be necessary to fully assess the generality of this conclusion.

A natural history of FUT2 polymorphism in humans

2009-06-01

Because pathogens are powerful selective agents, host cell surface molecules used by pathogens as identification signals can reveal the signature of selection. Most of them are oligosaccharides, synthesized by glycosyltransferases. One known example is balancing selection shaping ABO evolution as a consequence of both, A and B antigens being recognized as receptors by some pathogens, and anti-A and/or anti-B natural antibodies produced by hosts conferring protection against the numerous infectious agents expressing A and B motifs. These antigens can also be found in tissues other than blood if there is activity of another enzyme, FUT2, a fucosyltransferase responsible for ABO biosynthesis in body fluids. Homozygotes for null variants at this locus present the non-secretor phenotype (se), since they can not express ABO antigens in secretions. Multiple independent mutations have been shown to be responsible for the non-secretor phenotype, which is coexisting with the secretor phenotype in most populations. In this study, we have resequenced the coding region of FUT2 in 732 individuals from 39 worldwide human populations. We report a complex pattern of natural selection acting on the gene. While frequencies of secretor and non-secretor phenotypes are similar in different populations, the point mutations at the base of the phenotypes are different, with some variants showing a long history of balancing selection among Eurasian and African populations, and one recent variant showing a fast spread in East Asia, likely due to positive selection. Thus a convergent phenotype composition has been achieved through different mutations with different evolutionary histories.

Spatial inference of admixture proportions and secondary contact zones

Durand, E., Jay, F., Gaggiotti, O. E., Francois, O. — 2009-05-21

Genetic admixture of distinct gene pools is the consequence of complex spatio-temporal processes that could have involved massive migration and local mating during the history of a species. However current methods for estimating individual admixture proportions lack the incorporation of such a piece of information. Here, we extend Bayesian clustering algorithms by including global trend surfaces and spatial autocorrelation in the prior distribution on individual admixture coefficients. We test our algorithm by using spatially explicit and realistic coalescent simulations of colonization followed by secondary contact. By coupling our multiscale spatial analyses with a Bayesian evaluation of model complexity and fit, we show that the algorithm provides a correct description of smooth clinal variation, while still detecting zones of sharp variation when they are present in the data. We also apply our approach to understanding the population structure of the killifish, Fundulus heteroclitus, for which the algorithm uncovers a presumed contact zone in the Altantic coast of North-America.

Phylogenomics of C4 photosynthesis in sedges (Cyperaceae): multiple appearances and genetic convergence

2009-05-21

C₄ photosynthesis is an adaptive trait conferring an advantage in warm and open habitats. It originated multiple times and is presently reported in 18 plant families. It has been recently shown that phosphoenolpyruvate carboxylase (PEPC), a key enzyme of the C₄ pathway, evolved through numerous independent but convergent genetic changes in grasses (Poaceae). To compare the genetics of multiple C₄ origins on a broader scale, we reconstructed the evolutionary history of the C₄ pathway in sedges (Cyperaceae), the second most species-rich C₄ family. A sedge phylogeny based on two plastome genes (rbcL and ndhF) has previously identified six fully C₄ clades. A relaxed molecular clock was used to calibrate this tree and showed that the first C₄ acquisition occurred in this family between 19.6 and 10.1 million years ago. According to analyses of PEPC encoding genes (ppc), at least five distinct C₄ origins are present in sedges. Two C₄ Eleocharis species, which were unrelated in the plastid phylogeny, acquired their C₄-specific PEPC genes from a single source, probably through reticulate evolution or a horizontal transfer event. Acquisitions of C₄ PEPC in sedges have been driven by positive selection on at least 16 codons (3.5% of the studied gene segment). These sites underwent parallel genetic changes across the five sedge C₄ origins. Five of these sites underwent identical changes also in grass and eudicot C₄ lineages, indicating that genetic convergence is most important within families but that identical genetic changes occurred even among distantly related taxa. These lines of evidence give new insights into the constraints that govern molecular evolution, which are discussed here.

A recent adaptive transposable element insertion near highly conserved developmental loci in Drosophila melanogaster

Gonzalez, J., Macpherson, J. M., Petrov, D. A. — 2009-05-20

A recent genome-wide screen identified 13 transposable elements that are likely to have been adaptive during or after the spread of Drosophila melanogaster out of Africa. One of these insertions, Bari-Jheh, was associated with the selective sweep of its flanking neutral variation and with reduction of expression of one of its neighboring genes: Jheh3. Here, we provide further evidence that Bari-Jheh insertion is adaptive. We delimit the extent of the selective sweep and show that Bari-Jheh is the only mutation linked to the sweep. Bari-Jheh also lowers the expression of its other flanking gene, Jheh2. Subtle consequences of Bari-Jheh insertion on life history traits are consistent with the effects of reduced expression of the Jheh genes. Finally, we analyze molecular evolution of Jheh genes both in the long and the short term and conclude that Bari-Jheh appears to be a very rare adaptive event in the history of these genes. We discuss the implications of these findings for the detection and understanding of adaptation.

A machine-learning approach reveals that alignment properties alone can accurately predict inference of lateral gene transfer from discordant phylogenies

Roettger, M., Martin, W., Dagan, T. — 2009-05-14

Among the methods currently used in phylogenomic practice to detect the presence of lateral gene transfer (LGT), one of the most frequently employed is the comparison of gene tree topologies for different genes. In cases where the phylogenies for different genes are incompatible, or discordant, for well-supported branches there are three simple interpretations for the result: i) gene duplications (paralogy) followed by many independent gene losses has occurred, ii) LGT has occurred, or iii) the phylogeny is well-supported, but for reasons unknown is nonetheless incorrect. Here we focus on the third possibility by examining the properties of 22,437 published multiple sequence alignments, the Bayesian maximum likelihood trees for which either do or not suggest the occurrence of LGT by the criterion of discordant branches. The alignments that produce discordant phylogenies differ significantly in several salient alignment properties from those that do not. Using a support vector machine, we were able to predict the inference of discordant tree topologies with up to 80% accuracy from alignment properties alone.

A single-copy IS5-like transposon in the genome of a bdelloid rotifer

Gladyshev, E. A., Arkhipova, I. R. — 2009-05-14

In the course of sequencing telomeric chromosomal regions of the bdelloid rotifer Adineta vaga, we encountered an unusual DNA transposon. Unlike other bdelloid and, more generally, eukaryotic transposable elements (TEs), it exhibits similarity to prokaryotic insertion sequences (IS). Phylogenetic analysis indicates that this transposon, named IS5_Av, is related to the ISL2 group of the IS5 family of bacterial IS elements. Despite the apparent intactness of the single open reading frame coding for a DDE transposase and the perfect identity of its 213-bp terminal inverted repeats (TIRs), the element is present in only one copy per diploid genome. It does not exhibit any detectable levels of transcription, so that its transposase gene appears to be silent in the bdelloid host. While horizontal transfers of TEs between kingdoms are not known to happen in nature, it appears likely that IS5_Av underwent integration into the A. vaga genome relatively recently, but was not successful in adapting to the new host and failed to increase in copy number. Alternatively, it might be the only known member of a novel eukaryotic DNA TE superfamily which is so rare that its other members, if any, have not yet been identified in eukaryotic genomes sequenced to date.

Can GC content at third-codon positions be used as proxy for isochore composition?

Elhaik, E., Landan, G., Graur, D. — 2009-05-14

The isochore theory depicts the genomes of warm-blooded vertebrates as a mosaic of long genomic regions that are characterized by relatively homogeneous GC content. In the absence of genomic data, the GC content at third-codon positions of protein coding genes (GC3) was commonly used as proxy for the GC content of isochores. Oddly, in the post-genomic era, GC3 is still sometimes used as a proxy for the GC composition of isochores. Here, we use genic and genomic sequences from human, chimpanzee, cow, mouse, rat, chicken, and zebrafish to show that GC3 only explains a very small proportion of the variation in GC content of long genomic sequences flanking the genes (GCf), and what little correlation there is between GC3 and GCf was found to decay rapidly with distance from the gene. The coefficient of variation of GC3 was found to be much larger than that of GCf and, therefore, GC3 and GCf values are not comparable with each other. Comparisons of orthologous gene pairs from (1) human and chimpanzee and (2) mouse and rat show strong correlations between their GC3 values, but very weak correlations between their GCf values. We conclude that the GC content of third-codon position cannot be used as stand-in for isochoric composition.

A phylogenetic mixture model for gene family loss in parasitic bacteria

Spencer, M., Sangaralingam, A. — 2009-05-12

Gene families are frequently gained and lost from prokaryotic genomes. It is widely believed that the rate of loss was accelerated for some but not all gene families in lineages that became parasites or endosymbionts. This leads to a form of heterotachy which may be responsible for the poor performance of phylogeny estimation based on gene content. We describe a mixture model which accounts for this heterotachy. We show that this model fits data on the distribution of gene families across bacteria from the COG database much better than previous models. However, it still favours an artefactual tree topology in which parasites form a clade over the more plausible 16S topology. In contrast to a previous model of genome dynamics, our model suggests that the ancestral bacterium had a small genome. We suggest that models of gene family gain and loss are likely to be more useful for understanding genome dynamics than for estimating phylogenetic trees.

New Insights into Centromere Organization and Evolution from the White-cheeked Gibbon and Marmoset

2009-05-08

The evolutionary history of alpha-satellite DNA, the major component of primate centromeres, is hardly defined because of the difficulty in its sequence assembly and its rapid evolution when compared to most genomic sequences. By using several approaches, we have cloned, sequenced and characterized -satellite sequences from two species representing critical nodes in the primate phylogeny: the white-cheeked gibbon, a lesser ape and marmoset, a New World monkey. Sequence analyses demonstrate that white-cheeked gibbon and marmoset -satellite sequences are formed by units of ~171 bp and ~342 bp, respectively, and they both lack high-order structure found in humans and great apes. FISH characterization shows a broad dispersal of alpha-satellite in the white-cheeked gibbon genome including centromeric, telomeric and chromosomal interstitial localizations. On the other side centromeres in marmoset appear organized in highly divergent dimers roughly of 342bp that show a similarity between monomers much lower than previously reported dimers thus representing an ancient dimeric structure.

All these data shed light on the evolution of the centromeric sequences in Primates. Our results suggest radical differences in the structure, organization and evolution of -satellite DNA among different primate species, supporting the notion that i) all the centromeric sequence in Primates evolved by genomic amplification, unequal crossover and sequence homogenization using a 171bp monomer as basic seeding unit and ii) centromeric function is linked to relatively short repeated elements, more than higher-order structure.

Moreover, our data indicate that complex higher-order repeat structures are a peculiarity of the hominid lineage, showing the more complex organization in human.

A Comprehensive Overview of the Vertebrate p24 Family: Identification of a Novel Tissue-Specifically Expressed Member

Strating, J. R. P. M., van Bakel, N. N., Leunissen, J. A. M., Martens, G. J. M. — 2009-05-08

The members of the p24 protein family have an important but unclear role in transport processes in the early secretory pathway. The p24 family consists of four subfamilies (, β, and ), whereby the exact composition of the family varies among species. Despite more than 15 years of p24 research, the vertebrate p24 family is still surprisingly ill characterised. Here we describe the human, mouse, Xenopus and zebrafish orthologues of ten p24 family members and a new member that we term p24₅. Of these eleven p24 family members, nine are conserved throughout the vertebrate lineage, whereas two (p24₄ and p24₂) occur in some but not all vertebrates. We further show that all p24 proteins are widely expressed in mouse, except for p24₁ and p24₅ that display restricted expression patterns. Thus, we present for the first time a comprehensive overview of the phylogeny and expression of the vertebrate p24 protein family, including of a novel member.

INDELible: A Flexible Simulator of Biological Sequence Evolution

Fletcher, W., Yang, Z. — 2009-05-07

Many methods exist for reconstructing phylogenies from molecular sequence data, but few phylogenies are known and can be used to check their efficacy. Simulation remains the most important approach to testing the accuracy and robustness of phylogenetic inference methods. However, current simulation programs are limited, especially concerning realistic models for simulating insertions and deletions. We implement a portable and flexible application, named INDELible, for generating nucleotide, amino acid and codon sequence data by simulating insertions and deletions (indels) as well as substitutions. Indels are simulated under several models of indel length distribution. The program implements a rich repertoire of substitution models, including the general unrestricted model and non-stationary non-homogeneous models of nucleotide substitution, mixture and partition models that account for heterogeneity among sites, and codon models that allow the nonsynonymous/synonymous substitution rate ratio to vary among sites and branches. With its many unique features, INDELible should be useful for evaluating the performance of many inference methods, including those for multiple sequence alignment, phylogenetic tree inference and ancestral sequence or genome reconstruction.

Detecting ancient admixture and estimating demographic parameters in multiple human populations

Wall, J. D., Lohmueller, K. E., Plagnol, V. — 2009-05-06

We analyze patterns of genetic variation in extant human polymorphism data from the NIEHS SNPs project to estimate human demographic parameters. We update our previous work by considering a larger data set (more genes and more populations), and by explicitly estimating the amount of putative admixture between modern humans and archaic human groups (e.g., Neandertals, Homo erectus, H. floresiensis). We find evidence for this ancient admixture in European, East Asian and West African samples, suggesting that admixture between diverged hominin groups may be a general feature of recent human evolution.

Proteomics and comparative genomic investigations reveal heterogeneity in evolutionary rate of male reproductive proteins in mice (Mus domesticus)

2009-05-06

Male reproductive fitness is strongly affected by seminal fluid. In addition to interacting with the female environment, seminal fluid mediates important physiological characteristics of sperm, including capacitation and motility. In mammals, the male reproductive tract shows a striking degree of compartmentalization, with at least six distinct tissue types contributing material that is combined with sperm in an ejaculate. While studies of whole ejaculates have been undertaken in some species, we lack a comprehensive picture of the specific proteins produced by different accessory tissues. Here we perform proteomic investigations of six regions of the male reproductive tract in mice – seminal vesicles, anterior prostate, dorsolateral prostate, ventral prostate, bulbourethral gland, and bulbourethral diverticulum. We identify 766 proteins that could be mapped to 506 unique genes and compare them to a high-quality human seminal fluid dataset. We find that Gene Ontology functions of seminal proteins are largely conserved between mice and humans. By placing these data in an evolutionary framework, we show that seminal vesicle proteins have experienced a significantly higher rate of nonsynonymous substitution compared to the genome, which could be the result of adaptive evolution. In contrast, proteins from the other five tissues showed significantly lower nonsynonymous substitution, revealing a previously unappreciated level of evolutionary constraint acting on the majority of male reproductive proteins.

The evolutionary dynamics of autonomous non-ltr retrotransposons in the lizard Anolis carolinensis shows more similarity to fish than mammals

Novick, P. A., Basta, H., Floumanhaft, M., McClure, M. A., Boissinot, S. — 2009-05-06

The genome of the lizard Anolis carolinensis (the green anole) is the first non-avian reptilian genome sequenced. It offers a unique opportunity to comparatively examine the evolution of amniote genomes. We analyzed the abundance and diversity of non-LTR retrotransposons in the anole using the Genome Parsing Suite. We found that the anole genome contains an extraordinary diversity of elements. We identified 46 families of elements representing five clades (L1, L2, CR1, RTE and R4). Within most families, elements are very similar to each other suggesting that they have been inserted recently. The rarity of old elements suggests a high rate of turn-over, the insertion of new elements being offset by the loss of element-containing loci. Consequently, non-LTR retrotransposons accumulate in the anole at a low rate and are found in low copy number. This pattern of diversity shows some striking similarity with the genome of teleostean fish but contrasts greatly with the low-diversity and high copy number of mammalian L1 elements, suggesting a fundamental difference in the way mammals and non-mammalian vertebrates interact with their genomic parasites. The scarcity of divergent elements in anoles suggests that insertions have a deleterious effect and are eliminated by natural selection. We propose that the low abundance of non-LTR retrotransposons in the anole is related directly or indirectly to a higher rate of ectopic recombination in the anole relative to mammals.

Genetic admixture history of eastern Indonesia as revealed by Y-chromosome and mitochondrial DNA analysis

2009-05-04

Eastern Indonesia possesses more linguistic diversity than any other region in Southeast Asia, with both Austronesian (AN) languages that are of East Asian origin, as well as non-Austronesian (NAN) languages of likely Melanesian origin. Here, we investigated the genetic history of human populations from seven eastern Indonesian islands, including AN- and NAN-speakers, as well as the relationship between languages and genes, by means of non-recombining Y-chromosomal (NRY) and mitochondrial DNA (mtDNA) analysis. We found that the eastern Indonesian gene pool consists of East Asian as well as Melanesian components, as might be expected based on linguistic evidence, but also harbours putative indigenous eastern Indonesian signatures that perhaps reflect the initial occupation of the Wallacea by aboriginal hunter-gatherers already in Palaeolithic times. Furthermore, both NRY and mtDNA data showed a complete lack of correlation between linguistic and genetic relationships, most likely reflecting genetic admixture and/or language shift. In addition, we noted a small fraction of the NRY and mtDNA data shared between eastern Indonesians and Australian Aborigines likely reflecting an ancient link between Asia and Australia. Our data thus provide insights into the complex genetic ancestry history of eastern Indonesian islanders characterized by several admixture episodes, and demonstrate a clear example of the lack of the often-assumed correlation between the genes and languages of human populations.

Gene conversion maintains non-functional transposable elements in an obligate mutualistic endosymbiont

Cordaux, R. — 2009-05-04

Long-term bacterial endosymbionts typically exhibit reduced genomes, lack genes encoding recombination functions and transposable elements, such as insertion sequences (IS). In sharp contrast, I found that IS constitute 2.4% of the genome of the obligate mutualistic endosymbiont Wolbachia wBm. Although no IS copy is transpositionaly functional, I show that IS persist in wBm because of frequent recombinational gene conversion homogenizing homologous IS sequences. These results not only indicate that there exists a functional recombination molecular machinery in wBm, but they also suggest that, by slowing down the rate of IS degradation and loss, gene conversion may represent a major force influencing reductive evolution in wBm.

Evolution of a novel carotenoid binding protein responsible for crustacean shell color

Wade, N. M., Tollenaere, A., Hall, M. R., Degnan, B. M. — 2009-05-04

Carotenoids are commonly used by disparate metazoans to produce external coloration, often in direct association with specific proteins. In one such example, crustacyanin (CRCN) and the carotenoid astaxanthin combine to form a multimeric protein complex that is critical for the array of external shell colors in clawed lobsters. Through a combined biochemical, molecular genetic and bioinformatic survey of the distribution of CRCN across the animal kingdom, we have found that CRCNs are restricted to, but widespread among, malacostracan crustaceans. These crustacean-specific genes separate into two distinct clades within the lipocalin protein superfamily. We show that CRCN differentially localizes to colored shell territories and the underlying epithelium in panulirid lobsters. Given the paramount importance of CRCN in crustacean shell colors and patterns and the critical role these play in survival, reproduction and communication, we submit that the origin of the CRCN gene family early in the evolution of malacostracan crustaceans significantly contributed to the success of this group of arthropods.

Genomic drift and evolution of microsatellite DNAs in human populations

Takezaki, N., Nei, M. — 2009-04-30

In recent years, copy number variation (CNV) of DNA segments has become a hot topic in the study of genetic variation, and a large amount of CNVs has been uncovered in human populations. The CNVs involving the smallest units of DNA segments are microsatellite DNAs, and the evolutionary change of microsatellite DNAs is believed to occur mostly by the increase or decrease of one repeat unit at a time in a more or less neutral fashion. If we note that eukaryotic genomes contain millions of microsatellite loci, this pattern of nucleotide change is expected to generate random changes of genome size, i.e., genomic drift, and will provide a neutral model of CNV evolution. We therefore investigated the amount of variation of the total number of repeats (TNR) per individual concerned with 145 microsatellite loci in three human populations, Africans, Europeans, and Asians. It was shown that the TNR follows the normal distribution in all three populations and that the extent of variation of TNR is more than 50% greater in Africans than in Europeans and Asians as expected from the hypothesis of African origin of modern humans. If we consider all microsatellite loci in the human genome and compute the variation of the total number of nucleotides involved (TNN), it is possible to study the contribution of microsatellite loci to the genome size variation. This study has shown that the genome sizes of human individuals are affected considerably by genomic drift of microsatellite DNA alone. This pattern of evolution is similar to that of olfactory receptor (OR) genes previously studied in human populations and support the idea that the number of OR genes has evolved in a more or less neutral fashion. However, this conclusion does not necessarily apply to the genome-wide CNVs of various DNA segments, and it appears that long variant DNA fragments are deleterious and under purifying selection.

Adaptive gene loss reflects differences in the visual ecology of basal vertebrates

Davies, W. L., Collin, S. P., Hunt, D. M. — 2009-04-27

The agnathans (lampreys and hagfishes) are representatives of the jawless fishes and constitute the first lineage of extant vertebrates to evolve within chordate phylogenetic history. Previously, we showed that the southern hemisphere pouched lamprey, Geotria australis, has the potential for pentachromacy with the expression of five visual pigment (opsin) genes (LWS, SWS1, SWS2, RhA, RhB) in five different cone-like photoreceptor types for life in a brightly-lit environment exposed to a broad-spectrum of light. In contrast, the northern hemisphere sea lamprey Petromyzon marinus dwells in a wide range of depths that are relatively deeper than the epipelagic waters inhabited by G. australis. Thus, the light levels of the habitat in which the sea lamprey resides are greatly diminished and different regions of the light spectrum are differentially absorbed. Therefore, the visual systems of these two species of lamprey consititutes a natural experiment in which to study the selection pressures underlying opsin gene expression and the evolution of colour discrimination. By analysing the opsin genes of P. marinus, we show the expression of two intact retinal opsins, RhA and LWS, which, when regenerated with 11-cis retinal, give peak spectral sensitivities (_max values) of 501 nm and 536 nm, respectively. In contrast to G. australis, the genome of P. marinus possesses remnants of SWS1 and SWS2 pseudogenes, which with the loss of RhB, suggests that P. marinus is a dichromat. Using site-directed mutagenesis, we show that a single amino acid substiution (Ser to Pro) at site 164 is responsible for a blue-shift of 19 nm of the LWS visual pigment of P. marinus compared to G. australis, which may reflect habitat differences between the two species. Based on these studies, we propose that gene loss (or duplication) and subsequent mutation plays an important role in the evolution of colour vision and that the complement and tuning of these visual pigments reflect the ecology and light environment of these phylogenetically basal vertebrates.

Positive selection has driven the evolution of the Drosophila Insulin-like Receptor (InR) at different time scales

Guirao-Rico, S., Aguade, M. — 2009-04-27

The highly conserved insulin signaling pathway influences very diverse processes including intermediary metabolism, reproduction, aging and growth. The first pathway component is the insulin receptor that upon insulin binding triggers the signal-transduction cascade. Its variation, like that of other pathway components, might affect therefore many organismal traits. Variation at the Insulin-like receptor (InR) gene was surveyed both within Drosophila melanogaster and between species across the Drosophila phylogeny. In D. melanogaster, the level and pattern of variation at the ~8-Kb region surveyed did not provide any indication of a recent selective event in this region. Maximum likelihood (ML) analyses revealed the past action of purifying selection acting differentially both across the phylogeny and along the studied gene. Moreover, the ML analyses and the McDonald and Kreitman test revealed the footprint of positive selection driving amino acid changes to fixation in the branch separating the Sophophora and the Drosophila subgenera, and in the D. melanogaster lineage, respectively. The oldest selective events could have affected either the insulin binding or the signal transduction capacities of the receptor, whereas mutations affecting signal transduction would seem to underlie the more recent events.

Rye Bs disclose ancestral sequences in cereal genomes with a potential role on gametophyte chromatid segregation

Pereira, H. S., Barao, A., Caperta, A., Rocha, J., Viegas, W., Delgado, M. — 2009-04-24

Two sequence families, E3900 and D1100, are amplified on the subtelomeric domain of the long arm of rye B chromosomes, the region that controls its drive mechanism. In this work, PCR with a number of primers spanning E3900 show that the organization and nucleotide sequence of E3900 related portions are present and highly conserved on rye A chromosomes as well as in other cereals. Quantitative Real Time-PCR estimates two E3900 sequences to be represented in 100-150 copies on Bs, and at least as single copies on As. A novel E3900 related sequence, with a deletion that results in a frameshift and subsequently an open reading frame with putative DNA binding motifs is identified. Expression analysis of E3900 indicates identical transcription levels in leaves from plants with and without Bs, showing that the expression of these sequences must be silenced on Bs and tightly regulated on As in leaves. In contrast, E3900 transcription is up-regulated during meiosis exclusively in plants with Bs, maintaining a high level of transcription in the gametophyte. Interestingly, Bs not only influence their own chromatid segregation but also that of the regular chromosome complement of both rye and wheat. There is a drastic increase in frequency of disrupted metaphase and anaphase cells in the first mitosis of pollen grains carrying Bs, which appears to be due to anomalous adherences between sister chromatids. Taken together, this work provides insight into how E3900 sequences are potentially associated to important evolutionary mechanisms involved in basic cellular processes.

Diversity in degrees of freedom of mitochondrial transit peptides

Staiger, C., Hinneburg, A., Klosgen, R. B. — 2009-04-22

Most mitochondrial proteins are synthesized in the cytosol of eukaryotic cells as precursor proteins carrying N-terminal extensions called transit peptides or presequences which mediate their specific transport into mitochondria. However, plant cells possess a second potential target organelle for such transit peptides, the chloroplast. It can therefore be assumed that mitochondrial transit peptides in plants are exposed to an increased demand of specificity, which in turn leads to reduced degrees of freedom in these transit peptides compared to those of non-plant organisms. Our study investigates this hypothesis using fractal dimension. Statistical analysis of sequence data shows that the fractal dimension of mitochondrial transit peptides in plants is indeed significantly lower than that from non-plant organisms.

Rapid evolution of immune proteins in social insects

2009-04-22

The existence of behavioral traits connected to defense against pathogens manifests the importance of pathogens in the evolution of social insects. However, very little is known about how pathogen pressure has affected the molecular evolution of genes involved in their innate immune system. We have studied the sequence evolution of several immune genes in ants and honeybees. The results show high rates of evolution in both ants and honeybees as measured by the ratio of amino acid changes to silent nucleotide changes, the ratio being clearly higher than in Drosophila immune genes or in non-immunity genes of bees. This conforms to our expectations based on high pathogen pressure in social insects. The codon-based likelihood method found clear evidence of positive selection only in one ant gene, even though positive selection has earlier been found in both ant and termite immune genes. There is now indication that selection on the amino acid composition of the immune-related genes has been an important part in the fight against pathogens by social insects. However, we cannot distinguish in all the cases whether the high observed d_N/d_S ratio results from positive selection within a restricted part of the studied genes or from relaxation of purifying selection associated to effective measures of behaviorally based colony-level defenses.

Discrete shoot and root stem cell-promoting WUS/WOX5 functions are an evolutionary innovation of angiosperms

Nardmann, J., Reisewitz, P., Werr, W. — 2009-04-22

The morphologically diverse bodies of seed plants comprising gymnosperms and angiosperms, which separated some 350 mya, grow by the activity of meristems containing stem cell niches. In the dicot model Arabidopsis thaliana, these are maintained by the stem cell-promoting functions of WUS and WOX5 in the shoot and the root, respectively. Both genes are members of the WUSCHEL-related homeobox (WOX) gene family, which has a monophyletic origin in green algae. The establishment of the WOX gene phylogeny from basal land plants through gymnosperms to basal and higher angiosperms reveals three major branches: a basal clade consisting of WOX13-related genes present in some green algae and throughout all land plant genomes, a second clade containing WOX8/9/11/12 homologues and a modern clade restricted to seed plants. The analysis of the origin of the modern branch in two basal angiosperms (Amborella trichopoda, Nymphaea jamesoniana) and three gymnosperms (Pinus sylvestris, Ginkgo biloba, Gnetum gnemon) shows that all members of the modern clade consistently found in mono- and dicots exist at the base of the angiosperm lineage, including WUS and WOX5 orthologues. In contrast, our analyses indentify a single WUS/WOX5 homologue in all three gymnosperm genomes, consistent with a monophyletic origin in the last common ancestor of gymno- and angiosperms. Phylogenetic data, WUS and WOX5-specific evolutionary signatures as well as the expression pattern and stem cell-promoting function of the single gymnosperm WUS/WOX5 pro-orthologue in Arabidopsis indicate a gene duplication event followed by sub-functionalisation at the base of angiosperms.

Dinoflagellate spliced leader RNA genes display a variety of sequences and genomic arrangements

Zhang, H., Campbell, D. A., Sturm, N. R., Lin, S. — 2009-04-22

Spliced leader (SL) trans-splicing is a common mRNA processing mechanism in dinoflagellates, in which a 22-nt sequence is transferred from the 5’-end of a small non-coding RNA, the SL RNA, to the 5’-end of mRNA molecules. While the SL RNA gene was initially shown to be organized as tandem repeats with transcripts of 50-60 nt, shorter than most of their counterparts in other organisms, other gene organizations and transcript lengths were reported subsequently. To address the evolutionary gradient of gene organization complexity, we thoroughly examined transcript and gene organization of the SL RNA in a phylogenetically and ecologically diverse group of dinoflagellates representing four Orders. All these dinoflagellates possessed SL RNA transcripts of 50-60 nt, although in one species additional transcripts of up to 92 nt were also detected. At the genomic level, various combinations of SL RNA and 5S rRNA tandem gene arrays, including SL RNA-only, 5S rRNA-only, and mixed SL RNA-5S rRNA clusters, were amplified by PCR for six dinoflagellates, containing intergenic spacers ranging from 88 bp to over 1.2 kb. Of these species, no SL RNA-5S rRNA gene cluster (SL-5S cluster) was detected in Prorocentrum minimum, and only K. brevis showed the U6 snRNA gene associated with these mixed arrays. The 5S rRNA-only array was also found in three dinoflagellates, along with two SL RNA-5S rRNA-adjacent arrangements found in two other species that could represent junctions. Two species contained multimeric SL exon repeats with no associated intron. These results suggest that 1) both the SL RNA tandem repeat and the SL-5S cluster genomic organizations are an "ancient" and widespread feature within the phylum of dinoflagellates, and 2) rampant genomic duplication and recombination are ongoing independently in each dinoflagellate lineage, giving rise to the highly complex and diversified genomic arrangements of the SL RNA gene, while conserving the length and structure of the functional SL RNA.

Evolution of ultrasmall spliceosomal introns in highly reduced nuclear genomes

Slamovits, C. H., Keeling, P. J. — 2009-04-20

Introns reduction and loss is a significant component of genome compaction in many eukaryotic lineages, including yeasts, microsporidia, and some nucleomorphs. Nucleomorphs are the extremely reduced relicts of algal endosymbiont nuclei found in two lineages, cryptomonads and chlorarachniophytes. In cryptomonads introns are rare, or even lost altogether. In contrast, the nucleomorph of the chlorarachniophyte Bigelowiella natans contains the smallest nuclear genome known, but paradoxically also retained over 800 tiny spliceosomal introns, ranging from 18 to 21 nucleotides in length. Since introns have not been described in any other chlorarachniophyte nucleomorph, we do not know when these introns were reduced or whether they have been lost in other lineages. To gain insight into the evolution of these unique introns, we sequenced more than 150 spliceosomal introns in the nucleomorph of the chlorarachniophyte Gymnochlora stellata and compared size distribution, sequence features and patterns of gain/loss. To clarify the possible relationship between intron size and splicing efficiency we also analysed the outcome of 580 splicing events. Overall, these data indicate that the radical intron size reduction took place in the ancestor of all extant chlorarachniophytes and that although most introns have been retained through this reductive process, intron loss has also occurred. We also show that intron size is not static, and splicing is not determined strictly by size, but that size does play a strong role in splicing efficiency, likely as part of a combination of sequence features and size.

Accurate estimation of gene evolutionary rates using XRATE, with an application to transmembrane proteins

Heger, A., Ponting, C. P., Holmes, I. — 2009-04-20

XRATE implements algorithms for comparative annotation, ancestral reconstruction, evolutionary rate estimation and simulation. Its modeling repertoire includes phylogenetic stochastic context-free grammars and phylo-HMMs. Following earlier tests of XRATE as a machine-learning tool suitable for alignment annotation, we now report the first tests of XRATE as a precise quantitative instrument for estimating evolutionary rates. We implement a codon model similar to that of Goldman & Yang (1994) and show that XRATE's parameter estimates are consistent with those of PAML. To demonstrate its utility, we apply the model to measure the difference in selective strength () between intracellular and secreted regions of type I transmembrane proteins. In 215 out of 303 instances a complex model with individual for each region provides a better fit to the data than the simpler single value model. Secreted portions of type I transmembrane proteins show an elevation in similar to that seen for secreted protein genes. Less stringent purifying selection is thus a general property of the extracellular milieu, rather than being specific to only soluble and secreted proteins.

ERAD-derived Pre-Protein Transport across the 2nd Outermost Plastid Membrane of Diatoms

Hempel, F., Bullmann, L., Lau, J., Zauner, S., Maier, U. G. — 2009-04-17

The diatom Phaeodactylum tricornutum harbours a plastid that is surrounded by four membranes and evolved by way of secondary endosymbiosis. Like land plants, most of its plastid proteins are encoded as pre-proteins on the nuclear genome of the host cell and are resultantly redirected into the organelle. Because two more membranes are present in diatoms than the one pair surrounding primary plastids, the targeting situation is obviously different and more complex. In this work, we focus on pre-protein transport across the second outermost plastid membrane – an issue that was experimentally inaccessible until now. We provide first indications that our hypothesis of an ERAD (ER-associated degradation)-derived pre-protein transport system might be correct. Our data demonstrate that the symbiont-specific Der1 proteins, sDer1-1 and sDer1-2, form an oligomeric complex within the second outermost membrane of the complex plastid. Moreover, we present first evidence that the complex interacts with transit peptides of pre-proteins being transported across this membrane into the periplastidal compartment, but not with transit peptides of stromal-targeted proteins. Thus, the sDer1-complex might have an additional role in discriminating pre-proteins that are transported across the two outermost membranes from pre-proteins directed across all four membranes of the complex plastid. Altogether, our studies of the symbiont-specific ERAD-like machinery of diatoms suggest that a pre-existing cellular machinery was recycled to fulfill a novel function during the transition of a former free-living eukaryote into a secondary endosymbiont.