Genomic evidence for role of inversion 3RP of Drosophila melanogaster in facilitating climate change adaptation

Chromosomal inversion polymorphisms are common in animals and plants, and recent models suggest that alternative arrangements spread by capturing different combinations of alleles acting additively or epistatically to favour local adaptation. It is also thought that inversions typically maintain favoured combinations for a long time by suppressing recombination between alternative chromosomal arrangements. Here, we consider patterns of linkage disequilibrium and genetic divergence in an old inversion polymorphism in Drosophila melanogaster (In(3R)Payne) known to be associated with climate change adaptation and a recent invasion event into Australia. We extracted, karyotyped and sequenced whole chromosomes from two Australian populations, so that changes in the arrangement of the alleles between geographically separated tropical and temperate areas could be compared. Chromosome‐wide linkage disequilibrium (LD) analysis revealed strong LD within the region spanned by In(3R)Payne. This genomic region also showed strong differentiation between the tropical and the temperate populations, but no differentiation between different karyotypes from the same population, after controlling for chromosomal arrangement. Patterns of differentiation across the chromosome arm and in gene ontologies were enhanced by the presence of the inversion. These data support the notion that inversions are strongly selected by bringing together combinations of genes, but it is still not clear if such combinations act additively or epistatically. Our data suggest that climatic adaptation through inversions can be dynamic, reflecting changes in the relative abundance of different forms of an inversion and ongoing evolution of allelic content within an inversion.     

How chromosomal rearrangements shape adaptation and speciation: Case studies in Drosophila pseudoobscura and its sibling species Drosophila persimilis

The gene arrangements of Drosophila have played a prominent role in the history of evolutionary biology from the original quantification of genetic diversity to current studies of the mechanisms for the origin and establishment of new inversion mutations within populations and their subsequent fixation between species supporting reproductive barriers. This review examines the genetic causes and consequences of inversions as recombination suppressors and the role that recombination suppression plays in establishing inversions in populations as they are involved in adaptation within heterogeneous environments. This often results in the formation of clines of gene arrangement frequencies among populations. Recombination suppression leads to the differentiation of the gene arrangements which may accelerate the accumulation of fixed genetic differences among populations. If these fixed mutations cause incompatibilities, then inversions pose important reproductive barriers between species. This review uses the evolution of inversions in Drosophila pseudoobscura and D. persimilis as a case study for how inversions originate, establish and contribute to the evolution of reproductive isolation.     

Allozyme divergence between different gene arrangements in Drosophila subobscura

Allozyme and chromosomal polymorphism were analysed simultaneously in three population samples of Drosophila subobscura from Scotland, Germany and Italy. The study revealed significant genic divergence between the various chromosomal gene arrangements for the loci Acph, Lap and Pep, which are located inside inversions. No difference was found with the loci Me and Odh, which are both on the same chromosome but proximal to the structurally polymorphic region. Samples of a given gene arrangement taken from the different localities are genetically uniform. The data indicate that the observed non-random associations between enzyme genes and chromosomal inversions are the result of inhibited recombination in heterokaryotypes and gradual differentiation of the separated gene pools, rather than selection and coadaptation.     

Chromosomal inversion polymorphism leads to extensive genetic structure: a multilocus survey in Drosophila subobscura

The adaptive character of inversion polymorphism in Drosophila subobscura is well established. The O(ST) and O(3+4) chromosomal arrangements of this species differ by two overlapping inversions that arose independently on O(3) chromosomes. Nucleotide variation in eight gene regions distributed along inversion O(3) was analyzed in 14 O(ST) and 14 O(3+4) lines. Levels of variation within arrangements were quite similar along the inversion. In addition, we detected (i) extensive genetic differentiation between arrangements in all regions, regardless of their distance to the inversion breakpoints; (ii) strong association between nucleotide variants and chromosomal arrangements; and (iii) high levels of linkage disequilibrium in intralocus and also in interlocus comparisons, extending over distances as great as approximately 4 Mb. These results are not consistent with the higher genetic exchange between chromosomal arrangements expected in the central part of an inversion from double-crossover events. Hence, double crossovers were not produced or, alternatively, recombinant chromosomes were eliminated by natural selection to maintain coadapted gene complexes. If the strong genetic differentiation detected along O(3) extends to other inversions, nucleotide variation would be highly structured not only in D. subobscura, but also in the genome of other species with a rich chromosomal polymorphism.     

STUDIES OF LINKAGE IN POPULATIONS. XIV. HISTORICAL CHANGES IN FREQUENCIES OF GENE ARRANGEMENTS AND ARRANGEMENT COMBINATIONS IN NATURAL POPULATIONS OF DROSOPHILA ROBUSTA

Abstract Data are reported showing large directional changes in the frequencies of some gene arrangements and arrangement combinations in certain natural populations of Drosophila robusta in the eastern United States. The changes involve the same X‐chromosomal inversion differences in two of the three localities studied and similar autosomal inversions in all three. These genetic changes provide a rare opportunity to observe evolutionary forces at work in nature. They are interpreted as being due to natural selection.     

Molecular population genetics of the rp49 gene region in different chromosomal inversions of Drosophila subobscura.

Nucleotide variation at the ribosomal protein 49 (rp49) gene region has been studied in 75 lines of Drosophila subobscura belonging to four chromosomal arrangements (Ost, O3+4, O3+4+8, and O3+4+23). The location of the rp49 gene region within the inversion loop differs among heterokaryotypes: it is very close to one of the breakpoints in heterozygotes involving Ost chromosomes, while it is in a more central position in all other heterokaryotypes. The distribution of nucleotide polymorphism in the different arrangements is consistent with a monophyletic origin of the inversions. The data also provide evidence that gene conversion and possibly double crossover are involved in shuffling nucleotide variation among gene arrangements. The analyses reveal that the level of genetic exchange is higher when the region is located in a more central position of the inverted fragment than when it is close to the breakpoints. The pairwise difference distributions as well as the negative values of Tajima''s and Fu and Li''s statistics further support the hypothesis that nucleotide variation within chromosomal arrangements still reflects expansion after the origin of the inversions. Under the expansion model, we have estimated the time of origin of the studied inversions.     

Tracking changes in chromosomal arrangements and their genetic content during adaptation

There is considerable evidence for an adaptive role of inversions, but how their genetic content evolves and affects the subsequent evolution of chromosomal polymorphism remains controversial. Here, we track how life‐history traits, chromosomal arrangements and 22 microsatellites, within and outside inversions, change in three replicated populations of Drosophila subobscura for 30 generations of laboratory evolution since founding from the wild. The dynamics of fitness‐related traits indicated adaptation to the new environment concomitant with directional evolution of chromosomal polymorphism. Evidence of selective changes in frequency of inversions was obtained for seven of 23 chromosomal arrangements, corroborating a role for inversions in adaptation. The evolution of linkage disequilibrium between some microsatellites and chromosomes suggested that adaptive changes in arrangements involved changes in their genetic content. Several microsatellite alleles increased in frequency more than expected by drift in targeted inversions in all replicate populations. In particular, there were signs of selection in the O₃₊₄ arrangement favouring a combination of alleles in two loci linked to the inversion and changing along with it, although the lack of linkage disequilibrium between these loci precludes epistatic selection. Seven other alleles increased in frequency within inversions more than expected by drift, but were not in linkage disequilibrium with them. Possibly these alleles were hitchhiking along with alleles under selection that were not specific to those inversions. Overall, the selection detected on the genetic content of inversions, despite limited coverage of the genome, suggests that genetic changes within inversions play an important role in adaptation.     

Geographic pattern of allozyme and inversion polymorphism on chromosome O of Drosophila subobscura and its evolutionary origin

The chromosome O of Drosophila subobscura was studied with respect to genetic variability at three enzyme loci (Odh, Me, and Lap-4) and with respect to inversion polymorphism. Population samples were taken from seven localities along a north-south gradient from Sweden and Scotland to Tunisia.The chromosomal analysis revealed clinal frequency changes for gene arrangements from north to south. With the enzyme loci Odh and Me allele frequencies are similar throughout the distribution range. Both loci are located outside the common inversion complex O₃₊₄. On the other hand, frequency changes parallel to those of the gene arrangements were observed for the alleles of the Lap-locus. Nonrandom associations between Lap-alleles and the superimposed gene arrangements O_ST, O₃₊₄, O₃₊₄₊₈, and O₃₊₄₊₂₃ were found. These gene arrangements differ from each other with respect to allele frequencies at the Lap-locus but for a given gene arrangement the relative frequencies of Lap-alleles remain relatively constant along the north-south gradient. Thus allele frequencies at the Lap-locus can be predicted from inversion frequencies.These observations can be interpreted in such a way that the pattern of allozyme variation within gene arrangements is due to founder effects caused by the unique origin of inversions. The gene blocks in the different inversions seem to represent more or less separated gene pools. In polymorphic populations the coexistence of genetically differentiated inversions presumably gives rise to heterotic interaction.     

STRUCTURE AND POPULATION GENETICS OF THE BREAKPOINTS OF A POLYMORPHIC INVERSION IN DROSOPHILA SUBOBSCURA

Drosophila subobscura is a paleartic species of the obscura group with a rich chromosomal polymorphism. To further our understanding on the origin of inversions and on how they regain variation, we have identified and sequenced the two breakpoints of a polymorphic inversion of D. subobscura—inversion 3 of the O chromosome—in a population sample. The breakpoints could be identified as two rather short fragments (～300 bp and 60 bp long) with no similarity to any known transposable element family or repetitive sequence. The presence of the ～300‐bp fragment at the two breakpoints of inverted chromosomes implies its duplication, an indication of the inversion origin via staggered double‐strand breaks. Present results and previous findings support that the mode of origin of inversions is neither related to the inversion age nor species‐group specific. The breakpoint regions do not consistently exhibit the lower level of variation within and stronger genetic differentiation between arrangements than more internal regions that would be expected, even in moderately small inversions, if gene conversion were greatly restricted at inversion breakpoints. Comparison of the proximal breakpoint region in species of the obscura group shows that this breakpoint lies in a small high‐turnover fragment within a long collinear region (～300 kb).     

The Genetic Content of Chromosomal Inversions across a Wide Latitudinal Gradient

There is increasing evidence regarding the role of chromosomal inversions in relevant biological processes such as local adaptation and speciation. A classic example of the adaptive role of chromosomal polymorphisms is given by the clines of inversion frequencies in Drosophila subobscura, repeatable across continents. Nevertheless, not much is known about the molecular variation associated with these polymorphisms. We characterized the genetic content of ca. 600 individuals from nine European populations following a latitudinal gradient by analysing 19 microsatellite loci from two autosomes (J and U) and the sex chromosome (A), taking into account their chromosomal inversions. Our results clearly demonstrate the molecular genetic uniformity within a given chromosomal inversion across a large latitudinal gradient, particularly from Groningen (Netherlands) in the north to Málaga (Spain) in the south, experiencing highly diverse environmental conditions. This low genetic differentiation within the same gene arrangement across the nine European populations is consistent with the local adaptation hypothesis for th evolutionof chromosomal polymorphisms. We also show the effective role of chromosomal inversions in maintaining different genetic pools within these inverted genomic regions even in the presence of high gene flow. Inversions represent thus an important barrier to gene flux and can help maintain specific allelic combinations with positive effects on fitness. Consistent patterns of microsatellite allele-inversion linkage disequilibrium particularly in loci within inversions were also observed. Finally, we identified areas within inversions presenting clinal variation that might be under selection.     

Contrasting Histories of Three Gene Regions Associated with In(3l)payne of Drosophila Melanogaster

In the present report, we studied nucleotide variation in three gene regions of Drosophila melanogaster, spanning >5 kb and showing different degrees of association with the cosmopolitan inversion In(3-L)Payne. The analysis of sequence variation in the regions surrounding the breakpoints and the heat shock 83 (Hsp83) gene locus, located close to the distal breakpoint, revealed the absence of shared polymorphisms and the presence of a number of fixed differences between arrangements, indicating absence of genetic exchange. In contrast, for the esterase-6 gene region, located in the center of the inversion, we observed the presence of shared polymorphisms between arrangements suggesting genetic exchange. In the regions close to the breakpoints, the common St arrangement is 10 times more polymorphic than inverted chromosomes. We propose that the lack of recombination between arrangements in these regions coupled with genetic hitchhiking is the best explanation for the low heterozygosity observed in inverted lines. Using the data for the breakpoints, we estimate that this inversion polymorphism is around 0.36 million yr old. Although it is widely accepted that inversions are examples of balanced polymorphisms, none of the current neutrality tests including our Monte Carlo simulations showed significant departure from neutral expectations.     

Transpecific polymorphisms in an inversion linked esterase locus in Drosophila buzzatii

Nucleotide variation was studied in a 1.1 kb section of the coding region of an Esterase gene (Est-A) that maps in the center of the segments rearranged by polymorphic inversions in the cactophilic Drosophila buzzatii. We examine 30 homozygous second-chromosome lines differing in gene arrangement and three D. koepferae isofemale lines as outgroups. Our data show that Est-A is a highly polymorphic gene at both synonymous and replacement sites. Significant departures from homogeneity in the distribution of the ratio of silent polymorphism to divergence predicted by the neutral theory reveals a local excess of silent polymorphism. This is consistent with the presence of two apparent narrow peaks of elevated silent polymorphism surrounding nonconservative amino acid substitutions. These polymorphisms as well as others at synonymous and nonsynonymous sites are shared with D. koepferae. We suggest that the presence of shared nucleotide polymorphisms is probably due to interspecific gene flow and/or balancing selection acting on replacement variants and/or to a decreased probability of loss of ancestral polymorphisms caused by linkage to an adaptive inversion polymorphism. Recurrent mutation and persistence of neutral ancestral polymorphisms cannot, however, be ruled out. The analysis of the distribution of nucleotide variation among the three chromosomal arrangements sampled reveals that derived arrangements (J and JZ(3)) are less polymorphic than the ancestral ST, and that the widely distributed ST and J arrangements are genetically differentiated. However, a significant number of polymorphisms are shared between arrangements, suggesting frequent exchange either from gene conversion or from double crossovers in heterokaryotypes. Finally, our present results in combination with data of sequence variation at the breakpoints of inversion J suggest that this old gene arrangement has risen in frequency in relatively recent times.     

Nucleotide variation at the yellow gene region is not reduced in Drosophila subobscura: a study in relation to chromosomal polymorphism

In contrast to Drosophila melanogaster and Drosophila simulans, the yellow (y) gene region of Drosophila subobscura is not located in a region with a strong reduction in recombination. In addition, this gene maps very close to the breakpoints of different inversions that segregate as polymorphic in natural populations of D. subobscura. Therefore, levels of variation at the y gene region in this species relative to those found in D. melanogaster and D. simulans may be affected not only by the change in the recombinational environment, but also by the presence of inversion polymorphism. To further investigate these aspects, an approximately 5.4-kb region of the A (=X) chromosome including the y gene was sequenced in 25 lines of D. subobscura and in the closely related species Drosophila madeirensis and Drosophila guanche. The D. subobscura lines studied differed in their A-chromosomal arrangements, A(st), A(2), and A(1). Unlike in D. melanogaster and D. simulans, levels of variation at the y gene region of D. subobscura are not reduced relative to those found at other genomic regions in the same species (rp49, Acp70A, and Acph-1). This result supports the effect of the change in the recombinational environment of a particular gene on the level of neutral variation. In addition, nucleotide variation is affected by chromosomal polymorphism. A strong genetic differentiation is detected between the A(1) arrangement and either A(st) or A(2), but not between A(st) and A(2). This result is consistent with the location of the y gene relative to the breakpoints of inversions A(1) and A(2). In addition, the pattern of nucleotide polymorphism in A(st)+A(2) and A(1) seems to point out that variation at the y gene region within these chromosomal classes is in the phase transient to equilibrium. The estimated ages of these arrangements assuming a star genealogy indicate that their origin cannot predate the D. madeirensis split. Therefore, the present results are consistent with a chromosomal phylogeny where Am(1), which is an arrangement present in D. madeirensis but absent in current populations of D. subobscura, would be the ancestral arrangement.     

Molecular population genetics of the OBP83 genomic region in Drosophila subobscura and D. guanche: contrasting the effects of natural selection and gene arrangement expansion in the patterns of nucleotide variation

Chromosomal inversion polymorphism play a major role in the evolutionary dynamics of populations and species because of their effects on the patterns of genetic variability in the genomic regions within inversions. Though there is compelling evidence for the adaptive character of chromosomal polymorphisms, the mechanisms responsible for their maintenance in natural populations is not fully understood. For this type of analysis, Drosophila subobscura is a good model species as it has a rich and extensively studied chromosomal inversion polymorphism system. Here, we examine the patterns of DNA variation in two natural populations segregating for chromosomal arrangements that differentially affect the surveyed genomic region; in particular, we analyse both nucleotide substitutions and insertion/deletion variations in the genomic region encompassing the odorant-binding protein genes Obp83a and Obp83b (Obp83 region). We show that the two main gene arrangements are genetically differentiated, but are consistent with a monophyletic origin of inversions. Nevertheless, these arrangements interchange some genetic information, likely by gene conversion. We also find that the frequency spectrum-based tests indicate that the pattern of nucleotide variation is not at equilibrium; this feature probably reflects the rapid increase in the frequency of the new gene arrangement promoted by positive selection (that is an adaptive change). Furthermore, a comparative analysis of polymorphism and divergence patterns reveals a relaxation of the functional constraints at the Obp83b gene, which might be associated with particular ecological or demographic features of the Canary island endemic species D. guanche     

Recombination Load in a Chromosomal Inversion Polymorphism of Drosophila subobscura

Chromosomal inversions suppress recombination in heterokaryotypes and may help to maintain positive epistatic interactions among groups of alleles at loci contained in the inversion. Here I evaluate the protective effect of inversions on recombination when different chromosomal segments, or even the whole chromosome O of Drosophila subobscura, can be effectively prevented from undergoing recombination in several naturally occurring heterokaryotypes. The fitness of flies made homozygous for recombinant chromosomes was generally lower when compared to their nonrecombinant counterparts, thus suggesting that segregating gene arrangements in this species hold together favorable combinations of alleles that interact epistatically.     

Genetic exchange versus genetic differentiation in a medium-sized inversion of Drosophila: the A2/Ast arrangements of Drosophila subobscura

Chromosomal inversion polymorphism affects nucleotide variation at loci associated with inversions. In Drosophila subobscura, a species with a rich chromosomal inversion polymorphism and the largest recombinational map so far reported in the Drosophila genus, extensive genetic structure of nucleotide variation was detected in the segment affected by the O(3) inversion, a moderately sized inversion at Muller's element E. Indeed, a strong genetic differentiation all over O(3) and no evidence of a higher genetic exchange in the center of the inversion than at breakpoints were detected. In order to ascertain, whether other polymorphic and differently sized inversions of D. subobscura also exhibited a strong genetic structure, nucleotide variation in 5 gene regions (P236, P275, P150, Sxl, and P125) located along the A(2) inversion was analyzed in A(st) and A(2) chromosomes of D. subobscura. A(2) is a medium-sized inversion at Muller's element A and forms a single inversion loop in heterokaryotypes. The lower level of variation in A(2) relative to A(st) and the significant excess of low-frequency variants at polymorphic sites indicate that nucleotide variation at A(2) is not at mutation-drift equilibrium. The closest region to an inversion breakpoint, P236, exhibits the highest level of genetic differentiation (F(ST)) and of linkage disequilibrium (LD) between arrangements and variants at nucleotide polymorphic sites. The remaining 4 regions show a higher level of genetic exchange between A(2) and A(st) chromosomes than P236, as revealed by F(ST) and LD estimates. However, significant genetic differentiation between the A(st) and A(2) arrangements was detected not only at P236 but also in the other 4 regions separated from the nearest breakpoint by 1.2-2.9 Mb. Therefore, the extent of genetic exchange between arrangements has not been high enough to homogenize nucleotide variation in the center of the A(2) inversion. A(2) can be considered a typical successful inversion of D. subobscura according to its relative length. Chromosomal inversion polymorphism of D. subobscura might thus cause the genome of this species to be highly structured and to harbor different gene pools that might contribute to maintain adaptations to particular environments.     

The evolutionary history of Drosophila buzzatii. XXXV. Inversion polymorphism and nucleotide variability in different regions of the second chromosome

Inversions are portions of a chromosome where the gene order is reversed relative to a standard reference orientation. Because of reduced levels of recombination in heterokaryotypes, inversions have a potentially important effect on patterns of nucleotide variability in those genomic regions close to, or included in, the inverted fragments. Here we report sequence variation at three anonymous regions (STSs) located at different positions in relation to second-chromosome inversion breakpoints in 29 isochromosomal lines derived from an Argentinean population of Drosophila buzzatii. In agreement with previous findings in Drosophila, gene flux (crossing over and/or gene conversion) between arrangements seems to appreciably increase as we approach the middle sections of inversion 2j, and patterns of nucleotide variability within, as well as genetic differentiation between chromosome arrangements, are comparable to those observed at the molecular marker outside the inverted fragments. On the other hand, nucleotide diversity near the proximal breakpoint of inversion 2j is reduced when contrasted with that found at the other regions, particularly in the case of derived inverted chromosomes. Using the data from the breakpoint, we estimate that the inversion polymorphism is approximately 1.63 N generations old, where N is the effective population size. An excess of low-frequency segregating polymorphisms is detected; mostly in the ancestral 2st arrangement and probably indicating a population expansion that predates the coalescent time of inversion 2j. Heterogeneity in mutation rates between the markers linked to the inversions may be sufficient to explain the different levels of nucleotide diversity observed. When considered in the context of other studies on patterns of variation relative to physical distance to inversion breakpoints, our data appear to be consistent with the conclusion that inversions are unlikely to be "long-lived" balanced polymorphisms.     

Genetic loads and coadaptivity of chromosomal inversions I. Inversion polymorphism and genetic load on chromosome O in a Drosophila subobscura population from Petnica

Inversion polymorphism on chromosome O and polymorphism for the viability of determining genes have been studied in a natural population of Drosophila subobscura from Petnica (Serbia). The range of inversion polymorphism and the abundance of particular gene arrangements in the study population agree with a general pattern of inversion polymorphism of D. subobscura in Europe. The data obtained on the amount of genetic loads show that the D. subobscura population from Petnica displays a moderate degree of that polymorphism, compared to the other studied populations of these species. Therefore, the D. subobscura population from Petnica could be tentatively classified as an ecologically central population. Examination association of chromosomal, thus, inversion polymorphism with gene polymorphism, in the form of genetic loads show that differences exist in the mean viability among certain gene arrangements. The distribution of deleterious genes among chromosome O gene arrangements were non-random.     

Interchromosomal exchange of genetic information between gene arrangements on the third chromosome of Drosophila pseudoobscura

During the last 60 years, the inversion polymorphism on the third chromosome of Drosophila pseudoobscura has become a case study of the evolution of linked blocks of genes, isolated from each other by the suppression of recombination in heterozygotes for different inversions. Due to its location within inverted regions in most gene arrangements, the amylase (Amy) gene region can be used to elucidate the molecular pattern of evolution in these inversions. We studied this region in the Tree Line phylad of gene arrangements, with regard to both restriction site polymorphisms (RSP) and nucleotide sequences. The analysis of restriction maps, encompassing 26 kb, corroborates the cytogenetic phylogeny established on the basis of inversion breakpoints. However, we found that the 2.7 kb of nucleotide sequences of the AmyI gene are identical in both Estes Park and Hidalgo arrangements, despite the fact that these inversions arose independently from Tree Line. These contrasting results suggest that a homogenizing force, most likely gene conversion, is able to bring about localized exchanges between otherwise isolated gene arrangements.