Evidence for positive selection in the gene fruitless in Anastrepha fruit flies

<p>Abstract</p> <p>Background</p> <p>Many genes involved in the sex determining cascade have indicated signals of positive selection and rapid evolution across different species. Even though <it>fruitless </it>is an important gene involved mostly in several aspects of male courtship behavior, the few studies so far have explained its high rates of evolution by relaxed selective constraints. This would indicate that a large portion of this gene has evolved neutrally, contrary to what has been observed for other genes in the sex cascade.</p> <p>Results</p> <p>Here we test whether the <it>fruitless </it>gene has evolved neutrally or under positive selection in species of <it>Anastrepha </it>(Tephritidae: Diptera) using two different approaches, a long-term evolutionary analysis and a populational genetic data analysis. The first analysis was performed by using sequences of three species of <it>Anastrepha </it>and sequences from several species of <it>Drosophila </it>using the ratio of nonsynonymous to synonymous rates of evolution in PAML, which revealed that the <it>fru </it>region here studied has evolved by positive selection. Using Bayes Empirical Bayes we estimated that 16 sites located in the connecting region of the <it>fruitless </it>gene were evolving under positive selection. We also investigated for signs of this positive selection using populational data from 50 specimens from three species of <it>Anastrepha </it>from different localities in Brazil. The use of standard tests of selection and a new test that compares patterns of differential survival between synonymous and nonsynonymous in evolutionary time also provide evidence of positive selection across species and of a selective sweep for one of the species investigated.</p> <p>Conclusions</p> <p>Our data indicate that the high diversification of <it>fru </it>connecting region in <it>Anastrepha </it>flies is due at least in part to positive selection, not merely as a consequence of relaxed selective constraint. These conclusions are based not only on the comparison of distantly related taxa that show long-term divergence time, but also on recently diverged lineages and suggest that episodes of adaptive evolution in <it>fru </it>may be related to sexual selection and/or conflict related to its involvement in male courtship behavior.</p

Haplotype network estimated by statistical parsimony (TCS ver 1.21).

<p>Thicker lines represent nonsynonymous substitutions and small circles stand for inferred haplotypes. Haplotypes were labeled sequentially and the subsequent letters represent the respective identified species. O – <i>A. obliqua</i>, F – <i>A. fraterculus</i>, S – <i>A. sororcula</i>, G – <i>A. grandis</i>, T – <i>A. striata</i>, B – <i>A. bistrigata</i>, E– <i>A. serpentina</i>. Letters near lines represent internal mutations labels.</p

Sliding window plots of the z-scores of radically changed properties showing regions under positive-destabilizing selection in (A) female and (B) male isoform.

<p>DM/OD1 – DNA-binding and dimerization domain. OD2 – dimerization domain 2. α₁, α₂ and α₃ – alpha-helices that compose the predicted UBA-like domain. α* - disordered C-terminal tail, proposed to fold as an alpha-helix. Dashed horizontal line indicates the Bonferroni corrected significant limit (z-score = 3.07, <i>p</i><0.05, male exon; z-score = 2.95, <i>p</i><0.05, female exon).</p

Unrooted phylogenetic trees estimated by maximum likelihood.

<p>A) Male isoform. B) Female isoform. The <i>foreground</i> branch is marked by an asterisk (*). The branch lengths are as nucleotide substitutions per nucleotide site.</p

Neutrality tests for <i>dsx</i> DM/OD1 and common region without DM/OD1 and OD2 domains.

<p>Significant tests after Dunn-Šidàk correction are given in bold.</p>**<p><i>p</i><0.01;</p>*<p><i>p</i><0.05; a 0.10><i>p</i>>0.05.</p

Contingency analysis of synonymous and nonsynonymous vs tip or interior position in the haplotype network.

<p>Fisher's Exact Test probability under the null hypothesis of homogeneity: 2.7 10⁻⁶.</p

Amino acid physicochemical properties under positive destabilizing and purifying selection in male and female isoforms of <i>doublesex</i>.

<p>Properties under positive destabilizing selection are boldfaced.</p>*<p>- <i>p</i><0.05;</p>**<p>- <i>p</i><0.01;</p>***<p>- <i>p</i><0.001.</p

Permutation analysis of synonymous and nonsynonymous mutations versus position in the haplotype network (tip or interior).

<p>Permutational probability: 0.0004.</p

Contingency analyses of how synonymous and nonsynonymous mutations within DM/OD1 and common region are distributed across the haplotype network position (tip or interior).

<p>Fisher's Exact Test probability under the null hypothesis of homogeneity: 0.0008.</p