Multiple and diverse structural changes affect the breakpoint regions of polymorphic inversions across the Drosophila genus

Chromosomal polymorphism is widespread in the Drosophila genus, with extensive evidence supporting its adaptive character in diverse species. Moreover, inversions are the major contributors to the genus chromosomal evolution. The molecular characterization of a reduced number of polymorphic inversion breakpoints in Drosophila melanogaster and Drosophila subobscura supports that their inversions would have mostly originated through a mechanism that generates duplications staggered double-strand breaks and has thus the potential to contribute to their adaptive character. There is also evidence for inversion breakpoint reuse at different time scales. Here, we have characterized the breakpoints of two inversions of D. subobscura O4 and O8 involved in complex arrangements that are frequent in the warm parts of the species distribution area. The duplications detected at their breakpoints are consistent with their origin through the staggered-break mechanism, which further supports it as the prevalent mechanism in D. subobscura. The comparative analysis of inversions breakpoint regions across the Drosophila genus has revealed several genes affected by multiple disruptions due not only to inversions but also to single-gene transpositions and duplications

The molecular characterization of fixed inversions breakpoints unveils the ancestral character of the Drosophila guanche chromosomal arrangements

Cytological studies revealed that the number of chromosomes and their organization varies across species. The increasing availability of whole genome sequences of multiple species across specific phylogenies has confirmed and greatly extended these cytological observations. In the Drosophila genus, the ancestral karyotype consists of five rod-like acrocentric chromosomes (Muller elements A to E) and one dot-like chromosome (element F), each exhibiting a generally conserved gene content. Chromosomal fusions and paracentric inversions are thus the major contributors, respectively, to chromosome number variation among species and to gene order variation within chromosomal element. The subobscura cluster of Drosophila consists in three species that retain the genus ancestral karyotype and differ by a reduced number of fixed inversions. Here, we have used cytological information and the D. guanche genome sequence to identify and molecularly characterize the breakpoints of inversions that became fixed since the D. guanche-D. subobscura split. Our results have led us to propose a modified version of the D. guanche cytological map of its X chromosome, and to establish that (i) most inversions became fixed in the D. subobscura lineage and (ii) the order in which the four X chromosome overlapping inversions occurred and became fixed

A new spontaneous chromosomal inversion in a classical laboratory strain of <em>Drosophila subobscura</em>

Drosophila subobscura stands out for its rich chromosomal polymorphism in natural populations. Krimbas (1992) reviewed up to 66 spontaneous chromosomal inversions that combined into 79 arrangements. Some of these inversions are common in the whole range of the species distribution, but others are only present either at low frequencies across the species distribution area or in a restricted geographical area

Characterization of dFOXO binding sites upstream of the Insulin Receptor P2 promoter across the Drosophila phylogeny

The insulin/TOR signal transduction pathway plays a critical role in determining such important traits as body and organ size, metabolic homeostasis and life span. Although this pathway is highly conserved across the animal kingdom, the affected traits can exhibit important differences even between closely related species. Evolutionary studies of regulatory regions require the reliable identification of transcription factor binding sites. Here we have focused on the Insulin Receptor (InR) expression from its P2 promoter in the Drosophila genus, which in D. melanogaster is up-regulated by hypophosphorylated Drosophila FOXO (dFOXO). We have finely characterized this transcription factor binding sites in vitro along the 1.3 kb region upstream of the InR P2 promoter in five Drosophila species. Moreover, we have tested the effect of mutations in the characterized dFOXO sites of D. melanogaster in transgenic flies. The number of experimentally established binding sites varies across the 1.3 kb region of any particular species, and their distribution also differs among species. In D. melanogaster, InR expression from P2 is differentially affected by dFOXO binding sites at the proximal and distal halves of the species 1.3 kb fragment. The observed uneven distribution of binding sites across this fragment might underlie their differential contribution to regulate InR transcription

Inversion evolutionary rates might limit the experimental identification of inversion breakpoints in non-model species

Chromosomal inversions are structural changes that alter gene order but generally not gene content in the affected region. In Drosophila, extensive cytological studies revealed the widespread character of inversion polymorphism, with evidence for its adaptive character. In Drosophila subobscura, polymorphism affects both its four large autosomal elements and its X (A) chromosome. The characterization of eight of these autosomal inversions breakpoints revealed that most of them originated through the staggered-breaks mechanism. Here, we have performed chromosomal walks to identify the breakpoints of two X-chromosome widely distributed inversions ¿A2 and A1¿ of D. subobscura. Inversion A2 is considered a warm-adapted arrangement that exhibits parallel latitudinal clines in the species ancestral distribution area and in both American subcontinents, whereas inversion A1 is only present in the Palearctic region where it presents an east-west cline. The duplication detected at the A2 inversion breakpoints is consistent with its origin by the staggered-breaks mechanism. Inversion A1 breakpoints could not be molecularly identified even though they could be narrowly delimited. This result points to chromosome walking limitations when using as a guide the genome of other species. Limitations stem from the rate of evolution by paracentric inversions, which in Drosophila is highest for the X chromosome

The origin of chromosomal inversions as a source of segmental duplications in the Sophophora subgenus of Drosophila

Chromosomal inversions can contribute to the adaptation of organisms to their environment by capturing particular advantageous allelic combinations of a set of genes included in the inverted fragment and also by advantageous functional changes due to the inversion process itself that might affect not only the expression of flanking genes but also their dose and structure. Of the two mechanisms originating inversions ectopic recombination, and staggered double-strand breaks and subsequent repair only the latter confers the inversion the potential to have dosage effects and/or to generate advantageous chimeric genes. In Drosophila subobscura, there is ample evidence for the adaptive character of its chromosomal polymorphism, with an important contribution of some warm-climate arrangements such as E1+2+9+12. Here, we have characterized the breakpoints of inversion E12 and established that it originated through the staggered-break mechanism like four of the five inversions of D. subobscura previously studied. This mechanism that also predominates in the D. melanogaster lineage might be prevalent in the Sophophora subgenus and contribute to the adaptive character of the polymorphic and fixed inversions of its species. Finally, we have shown that the D. subobscura inversion breakpoint regions have generally been disrupted by additional structural changes occurred at different time scales

An easy route to the massive karyotyping of complex chromosomal arrangements in Drosophila

Inversion polymorphism is widespread in the Drosophila genus as well as in other dipteran genera. The presence of polytene chromosomes in some insect organs and, thus, the possibility to observe the different arrangements generated by inversions through a microscope enhanced the cytological study of this structural polymorphism. In several Drosophila species, these studies provided evidence for the adaptive character of this polymorphism, which together with the standing interest to uncover targets of natural selection has led to a renewed interest for inversion polymorphism. Our recent molecular characterization of the breakpoint regions of five inversions of the E chromosome of D. subobscura has allowed us to design a PCR-based strategy to molecularly identify the different chromosomal arrangements and, most importantly, to determine the E chromosome karyotype of medium- and large-sized samples from natural populations. Individuals of a test sample that were both cytologically and molecularly karyotyped were used to establish the strategy that was subsequently applied to karyotype a larger sample. Our strategy has proved to be robust and time efficient, and it lays therefore the groundwork for future studies of the E chromosome structural polymorphism through space and time, and of its putative contribution to adaptation

The discovery, distribution and diversity of DNA viruses associated with Drosophila melanogaster in Europe

International audienceDrosophila melanogaster is an important model for antiviral immunity in arthropods, but very few DNA viruses have been described from the family Drosophilidae. This deficiency limits our opportunity to use natural host-pathogen combinations in experimental studies, and may bias our understanding of the Drosophila virome. Here we report fourteen DNA viruses detected in a metagenomic analysis of approximately 6500 pool-sequenced Drosophila, sampled from 47 European locations between 2014 and 2016. These include three new nudiviruses, a new and divergent entomopoxvirus, a virus related to Leptopilina boulardi filamentous virus, and a virus related to Musca domestica salivary gland hypertrophy virus. We also find an endogenous genomic copy of galbut virus, a dsRNA partitivirus, segregating at very low frequency. Remarkably, we find that Drosophila Vesanto virus, a small DNA virus previously described as a bidnavirus, may be composed of up to 12 segments and thus represent a new lineage of segmented DNA viruses. Two of the DNA viruses, Drosophila Kallithea nudivirus and Drosophila Vesanto virus are relatively common, found in 2% or more of wild flies. The others are rare, with many likely to be represented by a single infected fly. We find that virus prevalence in Europe reflects the prevalence seen in publicly-available datasets, with Drosophila Kallithea nudivirus and Drosophila Vesanto virus the only ones commonly detectable in public data from wild-caught flies and large population cages, and the other viruses being rare or absent. These analyses suggest that DNA viruses are at lower prevalence than RNA viruses in D. melanogaster, and may be less likely to persist in laboratory cultures. Our findings go some way to redressing an earlier bias toward RNA virus studies in Drosophila, and lay the foundation needed to harness the power of Drosophila as a model system for the study of DNA viruses

Genomic analysis of European Drosophila melanogaster populations reveals longitudinal structure, continent-wide selection, and previously unknown DNA viruses

Genetic variation is the fuel of evolution, with standing genetic variation especially important for short-term evolution and local adaptation. To date, studies of spatiotemporal patterns of genetic variation in natural populations have been challenging, as comprehensive sampling is logistically difficult, and sequencing of entire populations costly. Here, we address these issues using a collaborative approach, sequencing 48 pooled population samples from 32 locations, and perform the first continent-wide genomic analysis of genetic variation in European Drosophila melanogaster. Our analyses uncover longitudinal population structure, provide evidence for continent-wide selective sweeps, identify candidate genes for local climate adaptation, and document clines in chromosomal inversion and transposable element frequencies. We also characterize variation among populations in the composition of the fly microbiome, and identify five new DNA viruses in our samples.Publisher PDFPeer reviewe

Drosophila evolution over space and time (DEST):A new population genomics resource

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome datasets from natural populations of this species have been published over the last years. A major challenge is the integration of disparate datasets, often generated using different sequencing technologies and bioinformatic pipelines, which hampers our ability to address questions about the evolution of this species. Here we address these issues by developing a bioinformatics pipeline that maps pooled sequencing (Pool-Seq) reads from D. melanogaster to a hologenome consisting of fly and symbiont genomes and estimates allele frequencies using either a heuristic (PoolSNP) or a probabilistic variant caller (SNAPE-pooled). We use this pipeline to generate the largest data repository of genomic data available for D. melanogaster to date, encompassing 271 previously published and unpublished population samples from over 100 locations in > 20 countries on four continents. Several of these locations have been sampled at different seasons across multiple years. This dataset, which we call Drosophila Evolution over Space and Time (DEST), is coupled with sampling and environmental meta-data. A web-based genome browser and web portal provide easy access to the SNP dataset. We further provide guidelines on how to use Pool-Seq data for model-based demographic inference. Our aim is to provide this scalable platform as a community resource which can be easily extended via future efforts for an even more extensive cosmopolitan dataset. Our resource will enable population geneticists to analyze spatio-temporal genetic patterns and evolutionary dynamics of D. melanogaster populations in unprecedented detail.DrosEU is funded by a Special Topic Networks (STN) grant from the European Society for Evolutionary Biology (ESEB). MK (M. Kapun) was supported by the Austrian Science Foundation (grant no. FWF P32275); JG by the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (H2020-ERC-2014-CoG-647900) and by the Spanish Ministry of Science and Innovation (BFU-2011-24397); TF by the Swiss National Science Foundation (SNSF grants PP00P3_133641, PP00P3_165836, and 31003A_182262) and a Mercator Fellowship from the German Research Foundation (DFG), held as a EvoPAD Visiting Professor at the Institute for Evolution and Biodiversity, University of Münster; AOB by the National Institutes of Health (R35 GM119686); MK (M. Kankare) by Academy of Finland grant 322980; VL by Danish Natural Science Research Council (FNU) grant 4002-00113B; FS Deutsche Forschungsgemeinschaft (DFG) grant STA1154/4-1, Project 408908608; JP by the Deutsche Forschungsgemeinschaft Projects 274388701 and 347368302; AU by FPI fellowship (BES-2012-052999); ET Israel Science Foundation (ISF) grant 1737/17; MSV, MSR and MJ by a grant from the Ministry of Education, Science and Technological Development of the Republic of Serbia (451-03-68/2020-14/200178); AP, KE and MT by a grant from the Ministry of Education, Science and Technological Development of the Republic of Serbia (451-03-68/2020-14/200007); and TM NSERC grant RGPIN-2018-05551.Peer reviewe