Obscura Species Group Research Articles

Response of fruit fly (Drosophila pseudoobscura) to diet manipulation of nutrient density

AbstractCaloric intake can greatly affect many aspects of an organism's life. A deficiency of calories can lead to stress resulting in decreased fecundity, insufficient calories to maintain tissues and increased lifespan. Conversely, increasing caloric density increases fecundity and decreases lifespan. Despite decades of work exploring food quality and quantity on physiology in the model species Drosophila melanogaster Meigan 1830 (Diptera: Drosophilidae) and the melanogaster group in general, relatively little work explores the physiological responses to diet manipulation in other Drosophila species, like the obscura species group. Here, we looked at the effects of five different caloric densities (0.5×, 0.75×, 1.0×, 1.5× and 3.0×) on food intake, body weight, body fat, fecundity and longevity in D. pseudoobscura Frolova & Astaurov, 1929 (Diptera: Drosophilidae). Comparing longevity and fecundity across diets, we found that heavy caloric concentration (3.0×) decreases lifespan and that calorie restriction (0.5× and 0.75×) led to significant decreases in fecundity and body weight. However, calorie concentration did not significantly increase D. pseudoobscura body fat. By expanding our understanding of the physiological responses to diet stress to D. pseudoobscura, we establish the framework for comparative work across Drosophila species. With this information, we can then identify which physiological responses to diet manipulation might be most conserved and comparable across species.

The Drosophila melanogaster Y-linked gene, WDY, is required for sperm to swim in the female reproductive tract

Unique patterns of inheritance and selection on Y chromosomes have led to the evolution of specialized gene functions. We report CRISPR mutants in Drosophila of the Y-linked gene, WDY, which is required for male fertility. We demonstrate that the sperm tails of WDY mutants beat approximately half as fast as those of wild-type and that mutant sperm do not propel themselves within the male ejaculatory duct or female reproductive tract. Therefore, although mature sperm are produced by WDY mutant males, and are transferred to females, those sperm fail to enter the female sperm storage organs. We report genotype-dependent and regional differences in sperm motility that appear to break the correlation between sperm tail beating and propulsion. Furthermore, we identify a significant change in hydrophobicity at a residue at a putative calcium-binding site in WDY orthologs at the split between the melanogaster and obscura species groups, when WDY first became Y-linked. This suggests that a major functional change in WDY coincided with its appearance on the Y chromosome. Finally, we show that mutants for another Y-linked gene, PRY, also show a sperm storage defect that may explain their subfertility. Overall, we provide direct evidence for the long-held presumption that protein-coding genes on the Drosophila Y regulate sperm motility.

Mass spectrometric characterisation of the major peptides of the male ejaculatory duct, including a glycopeptide with an unusual zwitterionic glycosylation

Peptides present in the seminal fluid of Drosophila melanogaster can function as antimicrobial agents, enzyme inhibitors and as pheromones that elicit physiological and behavioural responses in the post-mated female. Understanding the molecular interactions by which these peptides influence reproduction requires detailed knowledge of their molecular structures. However, this information is often lacking and cannot be gleaned from just gene sequences and standard proteomic data. We now report the native structures of four seminal fluid peptides (andropin, CG42782, Met75C and Acp54A1) from the ejaculatory duct of male D. melanogaster. The mature CG42782, Met75C and Acp54A1 peptides each have a cyclic structure formed by a disulfide bond, which will reduce conformational freedom and enhance metabolic stability. In addition, the presence of a penultimate Pro in CG42782 and Met75C will help prevent degradation by carboxypeptidases. Met75C has undergone more extensive post-translational modifications with the formation of an N-terminal pyroglutamyl residue and the attachment of a mucin-like O-glycan to the side chain of Thr4. Both of these modifications are expected to further enhance the stability of the secreted peptide. The glycan has a rare zwitterionic structure comprising an O-linked N-acetyl hexosamine, a hexose and, unusually, phosphoethanolamine. A survey of various genomes showed that andropin, CG42782, and Acp54A1 are relatively recent genes and are restricted to the melanogaster subgroup. Met75C, however, was also found in members of the obscura species groups and in Scaptodrosophila lebanonensis. Andropin is related to the cecropin gene family and probably arose by tandem gene duplication, whereas CG42782, Met75C and Acp54A1 possibly emerged de novo. We speculate that the post-translational modifications that we report for these gene products will be important not only for a biological function, but also for metabolic stability and might also facilitate transport across tissue barriers, such as the blood-brain barrier of the female insect. Biological significanceSeminal fluid peptides of D. melanogaster function as antimicrobials, enzyme inhibitors and as pheromones, eliciting physiological and behavioural responses in the post-mated female. A fuller understanding of how these peptides influence reproduction requires knowledge not only of their primary structure, but also of their post-translational modification. However, this information is often lacking and difficult to glean from standard proteomic data. The reported modifications, including the unusual glycosylation, adds much to our knowledge of this important class of peptides in this model organism, par excellence.

Complex Evolutionary History of the Y Chromosome in Flies of the Drosophila obscura Species Group.

The Drosophila obscura species group shows dramatic variation in karyotype, including transitions among sex chromosomes. Members of the affinis and pseudoobscura subgroups contain a neo-X chromosome (a fusion of the X with an autosome), and ancestral Y genes have become autosomal in species harboring the neo-X. Detailed analysis of species in the pseudoobscura subgroup revealed that ancestral Y genes became autosomal through a translocation to the small dot chromosome. Here, we show that the Y-dot translocation is restricted to the pseudoobscura subgroup, and translocation of ancestral Y genes in the affinis subgroup likely followed a different route. We find that most ancestral Y genes have translocated to unique autosomal or X-linked locations in different taxa of the affinis subgroup, and we propose a dynamic model of sex chromosome formation and turnover in the obscura species group. Our results suggest that Y genes can find unique paths to escape unfavorable genomic environments that form after sex chromosome–autosome fusions.

Chromosome-Level Assembly of Drosophila bifasciata Reveals Important Karyotypic Transition of the X Chromosome

The Drosophila obscura species group is one of the most studied clades of Drosophila and harbors multiple distinct karyotypes. Here we present a de novo genome assembly and annotation of D. bifasciata, a species which represents an important subgroup for which no high-quality chromosome-level genome assembly currently exists. We combined long-read sequencing (Nanopore) and Hi-C scaffolding to achieve a highly contiguous genome assembly approximately 193 Mb in size, with repetitive elements constituting 30.1% of the total length. Drosophila bifasciata harbors four large metacentric chromosomes and the small dot, and our assembly contains each chromosome in a single scaffold, including the highly repetitive pericentromeres, which were largely composed of Jockey and Gypsy transposable elements. We annotated a total of 12,821 protein-coding genes and comparisons of synteny with D. athabasca orthologs show that the large metacentric pericentromeric regions of multiple chromosomes are conserved between these species. Importantly, Muller A (X chromosome) was found to be metacentric in D. bifasciata and the pericentromeric region appears homologous to the pericentromeric region of the fused Muller A-AD (XL and XR) of pseudoobscura/affinis subgroup species. Our finding suggests a metacentric ancestral X fused to a telocentric Muller D and created the large neo-X (Muller A-AD) chromosome ∼15 MYA. We also confirm the fusion of Muller C and D in D. bifasciata and show that it likely involved a centromere-centromere fusion.

Widespread gene duplication and adaptive evolution in the RNA interference pathways of the Drosophila obscura group

BackgroundRNA interference (RNAi) related pathways provide defense against viruses and transposable elements, and have been implicated in the suppression of meiotic drive elements. Genes in these pathways often exhibit high levels of adaptive substitution, and over longer timescales show gene duplication and loss—most likely as a consequence of their role in mediating conflict with these parasites. This is particularly striking for Argonaute 2 (Ago2), which is ancestrally the key effector of antiviral RNAi in insects, but has repeatedly formed new testis-specific duplicates in the recent history of the obscura species-group of Drosophila.ResultsHere we take advantage of publicly available genomic and transcriptomic data to identify six further RNAi-pathway genes that have duplicated in this clade of Drosophila, and examine their evolutionary history. As seen for Ago2, we observe high levels of adaptive amino-acid substitution and changes in sex-biased expression in many of the paralogs. However, our phylogenetic analysis suggests that co-duplications of the RNAi machinery were not synchronous, and our expression analysis fails to identify consistent male-specific expression.ConclusionsThese results confirm that RNAi genes, including genes of the antiviral and piRNA pathways, have undergone multiple independent duplications and that their history has been particularly labile within the obscura group. However, they also suggest that the selective pressures driving these changes have not been consistent, implying that more than one selective agent may be responsible.

Molecular population genetics of the Polycomb genes in Drosophila subobscura.

Polycomb group (PcG) proteins are important regulatory factors that modulate the chromatin state. They form protein complexes that repress gene expression by the introduction of posttranslational histone modifications. The study of PcG proteins divergence in Drosophila revealed signals of coevolution among them and an acceleration of the nonsynonymous evolutionary rate in the lineage ancestral to the obscura group species, mainly in subunits of the Pcl-PRC2 complex. Herein, we have studied the nucleotide polymorphism of PcG genes in a natural population of D. subobscura to detect whether natural selection has also modulated the evolution of these important regulatory genes in a more recent time scale. Results show that most genes are under the action of purifying selection and present a level and pattern of polymorphism consistent with predictions of the neutral model, the exceptions being Su(z)12 and Pho. MK tests indicate an accumulation of adaptive changes in the SU(Z)12 protein during the divergence of D. subobscura and D. guanche. In contrast, the HKA test shows a deficit of polymorphism at Pho. The most likely explanation for this reduced variation is the location of this gene in the dot-like chromosome and would indicate that this chromosome also has null or very low recombination in D. subobscura, as reported in D. melanogaster.

Adaptive selection and coevolution at the proteins of the Polycomb repressive complexes in Drosophila.

Polycomb group (PcG) proteins are important epigenetic regulatory proteins that modulate the chromatin state through posttranslational histone modifications. These interacting proteins form multimeric complexes that repress gene expression. Thus, PcG proteins are expected to evolve coordinately, which might be reflected in their phylogenetic trees by concordant episodes of positive selection and by a correlation in evolutionary rates. In order to detect these signals of coevolution, the molecular evolution of 17 genes encoding the subunits of five Polycomb repressive complexes has been analyzed in the Drosophila genus. The observed distribution of divergence differs substantially among and along proteins. Indeed, CAF1 is uniformly conserved, whereas only the established protein domains are conserved in other proteins, such as PHO, PHOL, PSC, PH-P and ASX. Moreover, regions with a low divergence not yet described as protein domains are present, for instance, in SFMBT and SU(Z)12. Maximum likelihood methods indicate an acceleration in the nonsynonymous substitution rate at the lineage ancestral to the obscura group species in most genes encoding subunits of the Pcl-PRC2 complex and in genes Sfmbt, Psc and Kdm2. These methods also allow inferring the action of positive selection in this lineage at genes E(z) and Sfmbt. Finally, the protein interaction network predicted from the complete proteomes of 12 Drosophila species using a coevolutionary approach shows two tight PcG clusters. These clusters include well-established binary interactions among PcG proteins as well as new putative interactions.

Simpler is better: fewer non‐target insects trapped with a four‐component chemical lure vs. a chemically more complex food‐type bait for Drosophila suzukii

AbstractBaits – fermented food products – are generally attractive to many types of insects, which makes it difficult to sort through non‐target insects to monitor a pest species of interest. We test the hypothesis that a chemically simpler and more defined attractant developed for a target insect is more specific and attracts fewer non‐target insects than a chemically more complex food‐type bait. A four‐component chemical lure isolated from a food bait and optimized for the spotted wing drosophila (SWD), Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), was compared to the original wine/vinegar bait to assess the relative responses of non‐target insects. In several field experiments in Washington State, USA, it was shown that numbers of pest muscid flies, cutworm and armyworm moths, and pest yellowjackets were reduced in traps baited with the chemical lure compared to the wine/vinegar bait. In other field experiments in the states of Washington, Oregon, and New York, numbers of non‐target drosophilid flies were also reduced in traps baited with the chemical lure relative to wine/vinegar bait. In Washington, numbers of Drosophila melanogaster Meigen and Drosophila obscura Fallen species groups and Drosophila immigrans Sturtevant were reduced in the chemical lure traps, whereas in New York, D. melanogaster and D. obscura species groups, D. immigrans, Drosophila putrida Sturtevant, Drosophila simulans Sturtevant, Drosophila tripunctata Loew, and Chymomyza spp. numbers were reduced. In Oregon, this same effect was observed with the D. melanogaster species group. Taken together, these results indicate that the four‐component SWD chemical lure will be more selective for SWD compared to fermentation baits, which should reduce time and cost involved in trapping in order to monitor SWD.

Microscale Laser Surgery Demonstrates the Grasping Function of the Male Sex Combs in Drosophila melanogaster and Drosophila bipectinata

AbstractMale secondary sexual traits of animals are richly diversified in form and complexity, yet there are many species in which their precise function remains unknown. Within the genus Drosophila, species belonging to the melanogaster and obscura species groups have evolved a remarkable variety of sex combs, male‐limited secondary sexual traits located on the tarsi of both front legs. Information concerning sex comb function is minimal or absent, except for D. melanogaster, where previous studies indicate that the sex combs are used for grasping the female prior to copulation. These studies, however, do not unambiguously demonstrate comb function, because it has not been possible to ascribe observed behavioral outcomes of the various comb manipulations to changes in the combs per se. We used microscale laser surgery to manipulate comb size in D. melanogaster and D. bipectinata, and tested the hypothesis that the sex combs function as grasping devices in courtship, making them essential for copulation to ensue. Results of high‐resolution behavioral analysis in small observation arenas demonstrated that in both species in which sex combs were surgically eliminated, males were unable to grasp, mount or copulate. The combless foretarsi of these altered males slipped off the end (D. melanogaster) and sides (D. bipectinata) of the female abdomen when courting males attempted to grasp. In most cases, males whose sex combs were reduced but not completely removed exhibited similar copulation probabilities as surgical control males, a result we demonstrated in observation chambers as well as under more ecologically realistic conditions inside population cages where males and females interacted on the surface of fruit substrates. Thus, the sex combs in D. melanogaster and D. bipectinata are grasping devices, essential for mounting and copulation.

Signatures of selection and sex-specific expression variation of a novel duplicate during the evolution of the Drosophila desaturase gene family

The tempo and mode of evolution of loci with a large effect on adaptation and reproductive isolation will influence the rate of evolutionary divergence and speciation. Desaturase loci are involved in key biochemical changes in long-chain fatty acids. In insects, these have been shown to influence adaptation to starvation or desiccation resistance and in some cases act as important pheromones. The desaturase gene family of Drosophila is known to have evolved by gene duplication and diversification, and at least one locus shows rapid evolution of sex-specific expression variation. Here, we examine the evolution of the gene family in species representing the Drosophila phylogeny. We find that the family includes more loci than have been previously described. Most are represented as single-copy loci, but we also find additional examples of duplications in loci which influence pheromone blends. Most loci show patterns of variation associated with purifying selection, but there are strong signatures of diversifying selection in new duplicates. In the case of a new duplicate of desat1 in the obscura group species, we show that strong selection on the coding sequence is associated with the evolution of sex-specific expression variation. It seems likely that both sexual selection and ecological adaptation have influenced the evolution of this gene family in Drosophila.

Discovery of three new species of Drosophila obscura species group (Diptera: Drosophilidae) from Mount Kinabalu in Borneo

AbstractWe describe three new species of the Drosophila obscura species group, D. hypercephala, D. hideakii and D. quadrangula Gao & Toda, spp. nov., all discovered from high altitudes (>1500 m) on Mt Kinabalu, Sabah, Malaysia. These are the southernmost distribution records of the D. obscura group in the Oriental Region. The three new species are each morphologically similar to different species of the Old World D. obscura species subgroup. This suggests that they are descendants of different founder species that independently colonized the tropics from the subtropical or temperate zones during the glacial ages. The new species D. hypercephala is hypercephalic not only in males but also, although less distinct, in females, unlike other hypercephalic drosophilid species in which only males are hypercephalic. This new finding of a hypercephalic species from the subgenus Sophophora, of which species are relatively easy to culture in the laboratory and include the model species such as D. melanogaster and D. pseudoobscura, will promote genetic analyses on the development and the evolution of hypercephaly.

Colonization of North America by Drosophila subobscura: Ecological analysis of three communities of drosophilids in California

We have carried out bimonthly collections of Drosophila species during a 16 month period in three localities of California (Eureka, Davis, and Gilroy). D. subobscura, which has colonized North America only a few years ago, is well established and has an annual abundance cycle similar to that observed in the Old World. The differences between the three localities account for 26.4% of the species diversity. Davis and Gilroy exhibit similarities attributable to their similar environmental conditions. Seasonality accounts for 46% and 50.4 % of the species diversity in Davis and Gilroy respectively, but only for 39.4% in Eureka. Differences between years explain only a small percentage of the total species diversity. A correspondence analysis of the association between species and months of collection shows ecological similarities between D. subobscura and the Nearctic members of the obscura species group.

Horizontal transmission versus vertical inheritance of P elements in Drosophila and Scaptomyza: has the M-type subfamily spread from East Asia?

Polymerase chain reaction (PCR) screening for P elements was carried out in 77 species with a primer set highly specific for the M-type subfamily. In the course of this search M-type elements were detected in 29 species: In the melanogaster (subgroups montium and rhopaloa) and obscura species groups of Drosophila (25 out of 71 species examined), and in the genus Scaptomyza (four out of six species). M-type elements are present in all species of the montium subgroup investigated so far (21), but occur only sporadically in other groups. Within the montium subgroup 20 species possess only incomplete copies, only one species (D. lacteicornis) harbours apparently full-sized elements. In contrast, outside the montium subgroup almost all species with M-type elements carry full-sized copies suggesting transpositional activity, at least in the recent past. The interior section of the full-sized M-type element of D. lacteicornis was partially sequenced (936 bp). In addition, the complete sequences of four internally deleted M-type elements of D. lacteicornis, D. rufa, D. quadraria, and D. triauraria were determined. Sequence comparisons (including sequence data from previous investigations) revealed striking discrepancies between P element phylogeny and the cladogenesis of their host species. Among several possible pathways for interspecific transfers of M-type elements, we favour the hypothesis assuming the invasion of Scaptomyza as well as the obscura group species of Drosophila via independent transmission routes originating from Asian species of the montium subgroup of Drosophila. The logical geographic scenario for these events would be East-Asia.

Rapid Evolution of Two Odorant-Binding Protein Genes, Obp57d and Obp57e, in the Drosophila melanogaster Species Group

Genes encoding odorant-binding protein (OBP) form a large family in an insect genome. Two OBP genes, Obp57d and Obp57e, were previously identified to be involved in host-plant recognition in Drosophila sechellia. Here, by comparing the genomic sequences at the Obp57d/e locus from 27 Drosophila species, we found large differences in gene number between species. Phylogenetic analysis revealed that Obp57d and Obp57e in the D. melanogaster species group arose by gene duplication of an ancestral OBP gene that remains single in the obscura species group. Further gain and loss of OBP genes were observed in several lineages in the melanogaster group. Site-specific analysis of evolutionary rate suggests that Obp57d and Obp57e have functionally diverged from each other. Thus, there are two classes of gene number differences in the Obp57d/e region: the difference of the genes that have functionally diverged from each other and the difference of the genes that appear to be functionally identical. Our analyses demonstrate that these two classes of differences can be distinguished by comparisons of many genomic sequences from closely related species.

WHAT USE IS AN INFERTILE SPERM? A COMPARATIVE STUDY OF SPERM-HETEROMORPHIC DROSOPHILA

Sperm size and number are important determinants of male reproductive success. The genus Drosophila exhibits a remarkable diversity of sperm production strategies, including the production of multiple sperm morphs by individual males, a phenomenon called sperm heteromorphism. Sperm-heteromorphic Drosophila species in the obscura group produce large numbers of infertile "parasperm" in addition to fertile eusperm. Parasperm have been hypothesized to perform a number of roles in place of fertilization, predominantly focused on their potential function in postcopulatory sexual selection. However, the evolutionary significance of parasperm remains unknown. Here we measured several male and female morphological, behavioral, and life-history traits in 13 obscura group species to test competing hypotheses of parasperm function using comparative methods. We found that parasperm size was unrelated to female reproductive tract morphology but was negatively related to our two indices of sperm competition, suggesting that postcopulatory sexual selection may indeed have shaped the evolution of parasperm. We also found abundant coevolution between male and female reproductive traits. Some of these relationships have been found in both sperm-monomorphic and sperm-heteromorphic taxa, but others are dissimilar. We discuss the significance of our results to the evolution of reproductive traits and the elusive function of Drosophila parasperm.

Molecular phylogeny of the Drosophila obscura species group, with emphasis on the Old World species

BackgroundSpecies of the Drosophila obscura species group (e.g., D. pseudoobscura, D. subobscura) have served as favorable models in evolutionary studies since the 1930's. Despite numbers of studies conducted with varied types of data, the basal phylogeny in this group is still controversial, presumably owing to not only the hypothetical 'rapid radiation' history of this group, but also limited taxon sampling from the Old World (esp. the Oriental and Afrotropical regions). Here we reconstruct the phylogeny of this group by using sequence data from 6 loci of 21 species (including 16 Old World ones) covering all the 6 subgroups of this group, estimate the divergence times among lineages, and statistically test the 'rapid radiation' hypothesis.ResultsPhylogenetic analyses indicate that each of the subobscura, sinobscura, affinis, and pseudoobscura subgroups is monophyletic. The subobscura and microlabis subgroups form the basal clade in the obscura group. Partial species of the obscura subgroup (the D. ambigua/D. obscura/D. tristis triad plus the D. subsilvestris/D. dianensis pair) forms a monophyletic group which appears to be most closely related to the sinobscura subgroup. The remaining basal relationships in the obscura group are not resolved by the present study. Divergence times on a ML tree based on mtDNA data are estimated with a calibration of 30–35 Mya for the divergence between the obscura and melanogaster groups. The result suggests that at least half of the current major lineages of the obscura group originated by the mid-Miocene time (~15 Mya), a time of the last developing and fragmentation of the temperate forest in North Hemisphere.ConclusionThe obscura group began to diversify rapidly before invading into the New World. The subobscura and microlabis subgroups form the basal clade in this group. The obscura subgroup is paraphyletic. Partial members of this subgroup (D. ambigua, D. obscura, D. tristis, D. subsilvestris, and D. dianensis) form a monophyletic group which appears to be most closely related to the sinobscura subgroup.

CHROMOSOMAL EVOLUTION OF ELEMENTS B AND C IN THE SOPHOPHORA SUBGENUS OF DROSOPHILA: EVOLUTIONARY RATE AND POLYMORPHISM

The locations of 77 markers along the chromosomal elements B (41 markers) and C (36 markers) of Drosophila subobscura, D. pseudoobscura, and D. melanogaster were obtained by in situ hybridization on polytene chromosomes. In comparisons between D. subobscura and D. pseudoobscura, 10 conserved segments (accounting for 32% of the chromosomal length) were detected on element B and eight (17% of the chromosomal length) on element C. The fixation rate of paracentric inversions inferred by a maximum likelihood approach differs significantly between elements. Muller's element C (0.17 breakpoints/Mb/million years) is evolving two times faster than element B (0.08 breakpoints/Mb/million years). This difference in the evolutionary rate is paralleled by differences in the extent of chromosomal polymorphism in the corresponding lineages. Element C is highly polymorphic in D. subobscura, D. pseudoobscura, and in other obscura group species such as D. obscura and D. athabasca. In contrast, the level of polymorphism in element B is much lower in these species. The fixation rates of paracentric inversions estimated in the present study between species of the Sophophora subgenus are the highest estimates so far reported in the genus for the autosomes. At the subgenus level, there is also a parallelism between the high fixation rate and the classical observation that the species of the Sophophora subgenus tend to be more polymorphic than the species of the Drosophila subgenus. Therefore, the detected relationship between level of polymorphism and evolutionary rate might be a general characteristic of chromosomal evolution in the genus Drosophila.

CHROMOSOMAL EVOLUTION OF ELEMENTS B AND C IN THE SOPHOPHORA SUBGENUS OF DROSOPHILA: EVOLUTIONARY RATE AND POLYMORPHISM

The locations of 77 markers along the chromosomal elements B (41 markers) and C (36 markers) of Drosophila subobscura, D. pseudoobscura, and D. melanogaster were obtained by in situ hybridization on polytene chromosomes. In comparisons between D. subobscura and D. pseudoobscura, 10 conserved segments (accounting for 32% of the chromosomal length) were detected on element B and eight (17% of the chromosomal length) on element C. The fixation rate of paracentric inversions inferred by a maximum likelihood approach differs significantly between elements. Muller's element C (0.17 breakpoints/Mb/million years) is evolving two times faster than element B (0.08 breakpoints/Mb/million years). This difference in the evolutionary rate is paralleled by differences in the extent of chromosomal polymorphism in the corresponding lineages. Element C is highly polymorphic in D. subobscura, D. pseudoobscura, and in other obscura group species such as D. obscura and D. athabasca. In contrast, the level of polymorphism in element B is much lower in these species. The fixation rates of paracentric inversions estimated in the present study between species of the Sophophora subgenus are the highest estimates so far reported in the genus for the autosomes. At the subgenus level, there is also a parallelism between the high fixation rate and the classical observation that the species of the Sophophora subgenus tend to be more polymorphic than the species of the Drosophila subgenus. Therefore, the detected relationship between level of polymorphism and evolutionary rate might be a general characteristic of chromosomal evolution in the genus Drosophila.

Domesticated P Elements in the Drosophila montium Species Subgroup Have a New Function Related to a DNA Binding Property

Molecular domestication of a transposable element is defined as its functional recruitment by the host genome. To date, two independent events of molecular domestication of the P transposable element have been described: in the Drosophila obscura species group and in the Drosophila montium species subgroup. These P neogenes consist of stationary, nonrepeated sequences, potentially encoding 66-kDa repressor-like (RL) proteins. Here we investigate the function of the montium P neogenes. We provide evidence for the presence of RL proteins in two montium species (D. tsacasi and D. bocqueti) specifically expressed in adult and larval brain and gonads. We tested the hypothesis that the montium P neogenes' function is related to the repression of the transposition of distantly related mobile P elements which coexist in the genome. Our results strongly suggest that the montium P neogenes are not recruited to downregulate the P element transposition. Given that all the proteins encoded by mobile or stationary P homologous sequences show a strong conservation of the DNA binding domain, we tested the capacity of the RL proteins to bind DNA in vivo. Immunostaining of polytene chromosomes in D. melanogaster transgenic lines strongly suggests that montium P neogenes encode proteins that bind DNA in vivo. RL proteins show multiple binding to the chromosomes. We suggest that the property recruited in the case of the montium P neoproteins is their DNA binding property. The possible functions of these neogenes are discussed.