Molecular evolution of sex-biased genes in Drosophila

Studies of morphology, interspecific hybridization, protein/DNA sequences, and levels of gene expression have suggested that sex-related characters (particularly those involved in male reproduction) evolve rapidly relative to non-sex-related characters. Here we report a general comparison of evolutionary rates of sex-biased genes using data from cDNA microarray experiments and comparative genomic studies of Drosophila. Comparisons of nonsynonymous/synonymous substitution rates (d(N)/d(S)) between species of the D. melanogaster subgroup revealed that genes with male-biased expression had significantly faster rates of evolution than genes with female-biased or unbiased expression. The difference was caused primarily by a higher d(N) in the male-biased genes. The same pattern was observed for comparisons among more distantly related species. In comparisons between D. melanogaster and D. pseudoobscura, genes with highly biased male expression were significantly more divergent than genes with highly biased female expression. In many cases, orthologs of D. melanogaster male-biased genes could not be identified in D. pseudoobscura through a Blast search. In contrast to the male-biased genes, there was no clear evidence for accelerated rates of evolution in female-biased genes, and most comparisons indicated a reduced rate of evolution in female-biased genes relative to unbiased genes. Male-biased genes did not show an increased ratio of nonsynonymous/synonymous polymorphism within D. melanogaster, and comparisons of polymorphism/divergence ratios suggest that the rapid evolution of male-biased genes is caused by positive selection.     

The evolution of sex-biased genes and sex-biased gene expression

Differences between males and females in the optimal phenotype that is favoured by selection can be resolved by the evolution of differential gene expression in the two sexes. Microarray experiments have shown that such sex-biased gene expression is widespread across organisms and genomes. Sex-biased genes show unusually rapid sequence evolution, are often labile in their pattern of expression, and are non-randomly distributed in the genome. Here we discuss the characteristics and expression of sex-biased genes, and the selective forces that shape this previously unappreciated source of phenotypic diversity. Sex-biased gene expression has implications beyond just evolutionary biology, including for medical genetics.     

Molecular evolution of sex-biased genes in the Drosophila ananassae subgroup

Background  

Genes with sex-biased expression often show rapid molecular evolution between species. Previous population genetic and comparative genomic studies of Drosophila melanogaster and D. simulans revealed that male-biased genes have especially high rates of adaptive evolution. To test if this is also the case for other lineages within the melanogaster group, we investigated gene expression in D. ananassae, a species that occurs in structured populations in tropical and subtropical regions. We used custom-made microarrays and published microarray data to characterize the sex-biased expression of 129 D. ananassae genes whose D. melanogaster orthologs had been classified previously as male-biased, female-biased, or unbiased in their expression and had been studied extensively at the population-genetic level. For 43 of these genes we surveyed DNA sequence polymorphism in a natural population of D. ananassae and determined divergence to the sister species D. atripex and D. phaeopleura.     

Effects of X-linkage and sex-biased gene expression on the rate of adaptive protein evolution in Drosophila

Patterns of polymorphism and divergence in Drosophila protein-coding genes suggest that a considerable fraction of amino acid differences between species can be attributed to positive selection and that genes with sex-biased expression, that is, those expressed predominantly in one sex, have especially high rates of adaptive evolution. Previous studies, however, have been restricted to autosomal sex-biased genes and, thus, do not provide a complete picture of the evolutionary forces acting on sex-biased genes across the genome. To determine the effects of X-linkage on sex-biased gene evolution, we surveyed DNA sequence polymorphism and divergence in 45 X-linked genes, including 17 with male-biased expression, 13 with female-biased expression, and 15 with equal expression in the 2 sexes. Using both single- and multilocus tests for selection, we found evidence for adaptive evolution in both groups of sex-biased genes. The signal of adaptive evolution was particularly strong for X-linked male-biased genes. A comparison with data from 91 autosomal genes revealed a "fast-X" effect, in which the rate of adaptive evolution was greater for X-linked than for autosomal genes. This effect was strongest for male-biased genes but could be seen in the other groups as well. A genome-wide analysis of coding sequence divergence that accounted for sex-biased expression also uncovered a fast-X effect for male-biased and unbiased genes, suggesting that recessive beneficial mutations play an important role in adaptation.     

Patterns of synonymous codon usage in Drosophila melanogaster genes with sex-biased expression

The nonrandom use of synonymous codons (codon bias) is a well-established phenomenon in Drosophila. Recent reports suggest that levels of codon bias differ among genes that are differentially expressed between the sexes, with male-expressed genes showing less codon bias than female-expressed genes. To examine the relationship between sex-biased gene expression and level of codon bias on a genomic scale, we surveyed synonymous codon usage in 7276 D. melanogaster genes that were classified as male-, female-, or non-sex-biased in their expression in microarray experiments. We found that male-biased genes have significantly less codon bias than both female- and non-sex-biased genes. This pattern holds for both germline and somatically expressed genes. Furthermore, we find a significantly negative correlation between level of codon bias and degree of sex-biased expression for male-biased genes. In contrast, female-biased genes do not differ from non-sex-biased genes in their level of codon bias and show a significantly positive correlation between codon bias and degree of sex-biased expression. These observations cannot be explained by differences in chromosomal distribution, mutational processes, recombinational environment, gene length, or absolute expression level among genes of the different expression classes. We propose that the observed codon bias differences result from differences in selection at synonymous and/or linked nonsynonymous sites between genes with male- and female-biased expression.     

The effects of sex-biased gene expression and X-linkage on rates of adaptive protein sequence evolution in Drosophila

A faster rate of adaptive evolution of X-linked genes compared with autosomal genes may be caused by the fixation of new recessive or partially recessive advantageous mutations (the Faster-X effect). This effect is expected to be largest for mutations that affect only male fitness and absent for mutations that affect only female fitness. We tested these predictions in Drosophila melanogaster by using genes with different levels of sex-biased expression and by estimating the extent of adaptive evolution of non-synonymous mutations from polymorphism and divergence data. We detected both a Faster-X effect and an effect of male-biased gene expression. There was no evidence for a strong association between the two effects—modest levels of male-biased gene expression increased the rate of adaptive evolution on both the autosomes and the X chromosome, but a Faster-X effect occurred for both unbiased genes and female-biased genes. The rate of genetic recombination did not influence the magnitude of the Faster-X effect, ruling out the possibility that it reflects less Hill–Robertson interference for X-linked genes.     

Patterns of evolution of genes disrupted in expression in Drosophila species hybrids

Divergence between species in regulatory pathways may contribute to hybrid incompatibilities such as sterility. Consistent with this idea, genes involved in male fertility often evolve faster than most other genes both in amino acid sequence and in expression. Previously, we identified a panel of male-specific genes under-expressed in sterile male hybrids of Drosophila simulans and D. mauritiana relative to pure species, and we showed that this under-expression is associated with infertility. In a preliminary effort to assess the generalities in the patterns of evolution of these genes, I examined patterns of mRNA expression in three of these genes in sterile F 1 hybrid males of D. pseudoobscura and D. persimilis . F 1 hybrid males bearing D. persimilis X chromosomes under-expressed all these genes relative to the parental species, while hybrids bearing D. pseudoobscura X chromosomes under-expressed two of these three genes. Interestingly, the third gene, CG5762 , has undergone extensive amino acid evolution within the D. pseudoobscura species group, possibly driven by positive natural selection. We conclude that some of the same genes exhibit disruptions in expression within each of the two species groups, which could suggest commonalities in the regulatory architecture of sterility in these groups. Alternative explanations are also considered.     

Sebida: a database for the functional and evolutionary analysis of genes with sex-biased expression

We describe Sebida, a database of genes with sex-biased expression. The database integrates results from multiple, independent microarray studies comparing male and female gene expression in Drosophila melanogaster, Drosophila simulans and Anopheles gambiae. Sebida uses standard nomenclature, which allows individual genes to be compared across different microarray platforms and to be queried by gene name, symbol, or annotation number. In addition to ratios of male/female expression for each gene, Sebida also contains information useful for evolutionary studies, such as local recombination rate, degree of codon bias and interspecific divergence at synonymous and non-synonymous sites. AVAILABILITY: Sebida can be accessed at http://www.sebida.de     

Rates of DNA sequence evolution are not sex-biased in Drosophila melanogaster and D. simulans

To determine whether male- or female-biased mutation rates have affected the molecular evolution of Drosophila melanogaster and D. simulans, we calculated the male-to-female ratio of germline cell divisions ([symbol: see text]) from germline generation data and the male-to-female ratio of mutation rate ([symbol: see text]) by comparing chromosomal levels of nucleotide divergence. We found that the ratio of germline cell divisions changes from indicating a weak female bias to indicating a weak male bias as the age of reproduction increases. The range of [symbol: see text] values that we observed, however, does not lead us to expect much, if any, difference in mutation rate between the sexes. Silent and intron nucleotide divergence were compared between nine loci on the X chromosome and nine loci on the second and third chromosomes. The average levels of nucleotide divergence were not significantly different across the chromosomes, although both silent and intron sites show a trend toward slightly more divergence on the X. These results indicate a lack of sex- or chromosome-biased molecular evolution in D. melanogaster and D. simulans.      

Molecular evolution of Drosophila odorant receptor genes

A total of 752 odorant receptor (Or) genes, including pseudogenes, were identified in 11 Drosophila species and named after their orthologs in Drosophila melanogaster. The 813 Or genes, including 61 from D. melanogaster, were classified into 59 orthologous groups that are well supported by gene phylogeny. By reconciling with the gene family phylogeny, we estimated the number of gene duplication/loss events and intron gain/loss events in the species phylogeny. We found that these events are particularly frequent in Drosophila grimshawi, Drosophila willistoni, and obscura group. More than half of the duplicated genes stay as tandem arrays, whose size range from 2 to 8. These genes vary in sequence and some likely underwent positive selection, indicating that the gene duplication was important for flies to acquire new olfactory functions. We hypothesize that Or genes conferred the basic olfactory repertoire to ancestral flies before the speciation of the Drosophila and Sophophora subgenera about 40 Mya. This repertoire has been largely maintained in the current species, whereas lineage-specific gene duplication seems to have led to additional specialization in some species in response to specific ecological conditions.     

Rampant adaptive evolution in regions of proteins with unknown function in Drosophila simulans

Adaptive protein evolution is pervasive in Drosophila. Genomic studies, thus far, have analyzed each protein as a single entity. However, the targets of adaptive events may be localized to particular parts of proteins, such as protein domains or regions involved in protein folding. We compared the population genetic mechanisms driving sequence polymorphism and divergence in defined protein domains and non-domain regions. Interestingly, we find that non-domain regions of proteins are more frequent targets of directional selection. Protein domains are also evolving under directional selection, but appear to be under stronger purifying selection than non-domain regions. Non-domain regions of proteins clearly play a major role in adaptive protein evolution on a genomic scale and merit future investigations of their functional properties.     

Estimating the genomewide rate of adaptive protein evolution in Drosophila

When polymorphism and divergence data are available for multiple loci, extended forms of the McDonald-Kreitman test can be used to estimate the average proportion of the amino acid divergence due to adaptive evolution--a statistic denoted alpha. But such tests are subject to many biases. Most serious is the possibility that high estimates of alpha reflect demographic changes rather than adaptive substitution. Testing for between-locus variation in alpha is one possible way of distinguishing between demography and selection. However, such tests have yielded contradictory results, and their efficacy is unclear. Estimates of alpha from the same model organisms have also varied widely. This study clarifies the reasons for these discrepancies, identifying several method-specific biases in widely used estimators and assessing the power of the methods. As part of this process, a new maximum-likelihood estimator is introduced. This estimator is applied to a newly compiled data set of 115 genes from Drosophila simulans, each with each orthologs from D. melanogaster and D. yakuba. In this way, it is estimated that alpha approximately 0.4+/-0.1, a value that does not vary substantially between different loci or over different periods of divergence. The implications of these results are discussed.     

Molecular population genetics and evolution of Drosophila meiosis genes

While many functional elements of the meiotic process are well characterized in model organisms, the genetic basis of most of the natural phenotypic variation observed in meiotic pathways has not been determined. To begin to address this issue, we characterized patterns of polymorphism and divergence in the protein-coding regions of 33 genes across 31 lines of Drosophila melanogaster and 6 lines of Drosophila simulans. We sequenced genes known to be involved in chromosome segregation, recombination, DNA repair, and related heterochromatin binding. As expected, we found several of the genes to be highly conserved, consistent with purifying selection. However, a subset of genes showed patterns of polymorphism and divergence typical of other types of natural selection. Moreover, several intriguing differences between the two Drosophila lineages were evident: along the D. simulans lineage we consistently found evidence of adaptive protein evolution, whereas along the D. melanogaster lineage several loci exhibited patterns consistent with the maintenance of protein variation.     

Structural evolution of the Drosophila 5S ribosomal genes

We compare the 5S gene structure from nine Drosophila species. New sequence data (5S genes of D. melanogaster, D. mauritiana, D. sechellia, D. yakuba, D. erecta, D. orena, and D. takahashii) and already-published data (5S genes of D. melanogaster, D. simulans, and D. teissieri) are used in these comparisons. We show that four regions within the Drosophila 5S genes display distinct rates of evolution: the coding region (120 bp), the 5-flanking region (54–55 bp), the 3-flanking region (21–22 bp), and the internal spacer (149–206 bp). Intra- and interspecific heterogeneity is due mainly to insertions and deletions of 6–17-bp oligomers. These small rearrangements could be generated by fork slippages during replication and could produce rapid sequence divergence in a limited number of steps. Correspondence to: M. Wegnez     

Organization and expression of Drosophila tropomyosin genes

It has been shown (Jockusch &; Isenberg, 1981) that vinculin (130K protein) binds to actin and induces actin filaments to form bundles even at low ionic strength. Here, we present structural details on the vinculin molecule itself and on its interaction with actin. In negatively stained preparations, vinculin appeared as a globular protein with an average diameter of 85 Å. The ability of vinculin to form actin filament bundles was confirmed using shadowing techniques and gel analysis of sedimented material. Analysis of vinculin-induced paracrystals by optical diffraction and computer processing revealed their structural similarity to Mg-induced paracrystals. The lateral position of vinculin on surface-exposed actin filaments of such paracrystals was demonstrated directly in electron micrographs and indirectly by labelling vinculin with ferritin-coupled anti-vinculin F(ab′) fragments. Polymerization of actin in the presence of vinculin-coated polystyrene beads did not result in an “end-on” binding of filaments to the beads. Rather, actin bundles were laterally associated with the whole surface of the beads, from where they radiated in a star-like pattern. The growth of actin filaments onto myosin subfragment-I decorated, vinculin-incubated. fixed filament fragments was not inhibited, as was shown directly by electron microscopy and monitored viscometrically in a nucleation assay. These results suggest that in vivo at the site of an adhesion plaque vinculin may link actin filaments together into a suitable configuration to interact with the plasma membrane.     

The role of laterally transferred genes in adaptive evolution

Background

Bacterial genomes develop new mechanisms to tide them over the imposing conditions they encounter during the course of their evolution. Acquisition of new genes by lateral gene transfer may be one of the dominant ways of adaptation in bacterial genome evolution. Lateral gene transfer provides the bacterial genome with a new set of genes that help it to explore and adapt to new ecological niches.

Methods

A maximum likelihood analysis was done on the five sequenced corynebacterial genomes to model the rates of gene insertions/deletions at various depths of the phylogeny.

Results

The study shows that most of the laterally acquired genes are transient and the inferred rates of gene movement are higher on the external branches of the phylogeny and decrease as the phylogenetic depth increases. The newly acquired genes are under relaxed selection and evolve faster than their older counterparts. Analysis of some of the functionally characterised LGTs in each species has indicated that they may have a possible adaptive role.

Conclusion

The five Corynebacterial genomes sequenced to date have evolved by acquiring between 8 – 14% of their genomes by LGT and some of these genes may have a role in adaptation.

     

Adaptive evolution of newly emerged micro-RNA genes in Drosophila

How often micro-RNA (miRNA) genes emerged and how fast theyevolved soon after their emergence are some of the central questionsin the evolution of miRNAs. Because most known miRNA genes areancient and highly conserved, these questions can be best answeredby identifying newly emerged miRNA genes. Among the 78 miRNAgenes in Drosophila reported before 2007, only 5 are confirmedto be newly emerged in the genus (although many more can befound in the newly reported data set; e.g., Ruby et al. 2007;Stark et al. 2007; Lu et al. 2008). These new miRNA genes haveundergone numerous changes, even in the normally invariant maturesequences. Four of them (the miR-310/311/312/313 cluster, denotedmiR-310s) were duplicated from other conserved miRNA genes.The fifth one (miR-303) appears to be a very young gene, originatingde novo from a non-miRNA sequence recently. We sequenced these5 miRNA genes and their neighboring regions from a worldwidecollection of Drosophila melanogaster lines. The levels of divergenceand polymorphism in these miRNA genes, vis-à-vis thoseof the neighboring DNA sequences, suggest that these 5 genesare evolving adaptively. Furthermore, the polymorphism patternof miR-310s in D. melanogaster is indicative of hitchhikingunder positive selection. Thus, a large number of adaptive changesover a long period of time may be essential for the evolutionof newly emerged miRNA genes.     

Germline transformation with Drosophila mutant actin genes induces constitutive expression of heat shock genes

Two Drosophila mutants KM75 and HH5, which are mutated in the act88F actin gene specific for the indirect flight muscles (IFM), synthesize heat shock proteins (hsps) constitutively in a tissue-specific manner. We have introduced cloned mutant act88F genes into a strain containing the wild-type act88F allele by P-element-mediated transformation. Flies transformed with a 4.05 kb KM75 act88F gene fragment encoding the p42 actin variant express both p42 and hsps specifically in the IFM. Using normal/mutant chimeric genes, the mutation sites of KM75 and HH5 were mapped within the sequence encoding the last 72 amino acids of actin. An in vitro mutated gene encoding a protein that lacks the 72 carboxy-terminal amino acids also induces constitutive hsp synthesis.