Inbreeding reveals interpopulation differences in inversion polymorphism of Drosophila subobscura

The adaptive significance of inversion polymorphism of Drosophila subobscura is well established. However, little is known about gene combinations within inversions which are coadapted because of population-specific effects. We studied this aspect of Dobzhansky's coadaptation hypothesis, using the systematic inbreeding method. Differences in magnitude and quality of inversion polymorphism in two ecologically and topologically distinct habitats were compared after several generations of continuous full-sib inbreeding. Populations from the two habitats differ in frequency of homokaryotypes after third and fifth generation of inbreeding and in the levels of homozygosity of different gene arrangements. The effect of homozygosity appears population and chromosome specific.     

Ultrastructure of regions containing homologous loci in polytene chromosomes of Drosophila melanogaster and Drosophila subobscura

We have used a new approach involving in situ hybridisation and electron microscopy to establish ultrastructural homologies between polytene chromosome regions of Drosophila melanogaster and Drosophila subobscura. Twelve probes were chosen to cover all the chromosomal elements: the myospheroid gene, the collagen type IV gene, the collagen-like gene, the w26 homeobox gene, the β3 tubulin gene, the kinesin heavy chain gene, the tryptophan hydrolase gene, the Hsp82, Hsp22–26 and Hsp23–28, Hsp68, Hsp70 genes and the β unit of the F₀–F₁ ATPase gene. Most of these loci were previously undescribed in D. subobscura and imprecisely located in D. melanogaster. We have demonstrated here, by an ultrastructural analysis of each chromosomal region, that homologous genetic loci tend to show a similar ultrastructure in the two species. With a few exceptions, the structural homology extends to the chromosomal regions surrounding the loci. In some cases, however, no structurally recognisable homology can be seen either in the locus or in its flanking regions. Received: 15 December 1996; in revised form: 15 October 1997 / Accepted: 28 January 1998     

Viabilities of originally natural O-chromosomal inversion homo- and heterokaryotypes in Drosophila subobscura

The influence of epistatic interactions of lethal and non-lethal genes upon viability of Drosophila inversion karyotypes is poorly known. In this paper we present comparative results for viabilities of 21 originally natural O-inversion homo- and 38 heterokaryotypes in a D. subobscura population. We observed strong heterotic effect in viability of O-lethal heterozygotes irrespective of different inversion backgrounds, which indicates a mechanism for protection of a great number of lethal genes. In O-non-lethal heterozygotes the heterotic effect in viability was absent irrespective of different inversion backgrounds. Our results showed a great extent of genetic load and high abundance of O-chromosomal arrangements in the population analyzed. It belongs to the set of central European populations. An epistasis of lethal genes present in O-inversion hetero- and not present in O-inversion homokaryotypes of moderate or low frequencies could be good example for co-adaptation of chromosomal inversions with regard to the genetic load. This represents a more efficient mechanism for limitation of genetic load than alternative mechanisms for protection of lethals. Except for lethal genes, possible epistatic interactions of mildly deleterious (subvital) genes, could also be responsible for limiting the extent of genetic load in natural D. subobscura populations. We demonstrated a non-random distribution of several combinations of viability classes among different O-inversion homo- and heterokaryotypes. As a consequence of that, the viabilities of the O-inversion homokaryotypes compared to heterokaryotypes were significantly higher at low frequencies than in moderate or high frequencies. This shows frequency-dependence as a mechanism of balancing selection for protection of O-chromosomal inversions in natural D. subobscura populations. In addition, the viabilities of the O-inversion homokaryotypes of lower frequency, compared to homokaryotypes of moderate or high frequency, were significantly higher. This again indicates the existence of supergene selection as another mechanism for protection of chromosomal inversions, as co-adapted complexes in natural D. subobscura populations.     

The effect of different concentrations of lead on inversion polymorphism in Drosophila subobscura

Drosophila subobscura is a wild Drosophila species that is spread over almost all of Europe. It possesses an uniquely rich inversion polymorphism on all five long chromosomes. This polymorphism is to a certain degree associated with the variation and dynamics of ecological factors in space and time. We analyzed the changes of inversion polymorphism components of Drosophila subobscura flies maintained on media with different concentrations of lead in laboratory conditions. The effects of lead on inversion polymorphism were observed by cytological analysis of gene arrangements on all of the five acrocentric chromosomes, as well as by cytological analysis of karyotypes on all of the four autosomes. The frequencies of particular gene arrangements on the four autosomes changed significantly in the samples maintained on medium not supplemented with lead. The frequencies of some gene arrangements on all of the five acrocentric chromosomes changed significantly in the flies maintained on media supplemented with lead. The length of exposure to different lead concentrations results in a significant change in the frequency of a few gene arrangements on two autosomes. However, the results show that different concentrations of lead, as well as the length of exposure, do not affect major parameters of inversion polymorphism. The results suggest that some gene arrangements could be linked with adaptive processes in evolving heavy metal resistance.     

Localization and characterization of X chromosome inversion breakpoints separating Drosophila mojavensis and Drosophila arizonae

Ectopic exchange between transposable elements or other repetitive sequences along a chromosome can produce chromosomal inversions. As a result, genome sequence studies typically find sequence similarity between corresponding inversion breakpoint regions. Here, we identify and investigate the breakpoint regions of the X chromosome inversion distinguishing Drosophila mojavensis and Drosophila arizonae. We localize one inversion breakpoint to 13.7 kb and localize the other to a 1-Mb interval. Using this localization and assuming microsynteny between Drosophila melanogaster and D. arizonae, we pinpoint likely positions of the inversion breakpoints to windows of less than 3000 bp. These breakpoints define the size of the inversion to approximately 11 Mb. However, in contrast to many other studies, we fail to find significant sequence similarity between the 2 breakpoint regions. The localization of these inversion breakpoints will facilitate future genetic and molecular evolutionary studies in this species group, an emerging model system for ecological genetics.     

The role of genic selection in the establishment of inversion polymorphism in Drosophila subobscura

A model for explaining the establishment of newly arisen inversions in natural populations, in which the inverted segment may be selected for if its load of deleterious mutations is smaller than the average load of the noninverted segment in the population, is tested for Drosophila subobscura. The results show that for new inversions, originally with no deleterious alleles, the expected cumulative distribution of inversion lengths fits fairly well with the observed one. Therefore, genic selection may be an important cause of the establishment of newly arisen inversions in natural populations of D. subobscura. The applicability of the model to the maintenance of the inversion polymorphism present in this species; is discussed.     

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.     

Evolutionary domestication in Drosophila subobscura

The domestication of plants and animals is historically one of the most important topics in evolutionary biology. The evolutionary genetic changes arising from human cultivation are complex because of the effects of such varied processes as continuing natural selection, artificial selection, deliberate inbreeding, genetic drift and hybridization of different lineages. Despite the interest of domestication as an evolutionary process, few studies of multicellular sexual species have approached this topic using well-replicated experiments. Here we present a comprehensive study in which replicated evolutionary trajectories from several Drosophila subobscura populations provide a detailed view of the evolutionary dynamics of domestication in an outbreeding animal species. Our results show a clear evolutionary response in fecundity traits, but no clear pattern for adult starvation resistance and juvenile traits such as development time and viability. These results supply new perspectives on the confounding of adaptation with other evolutionary mechanisms in the process of domestication.     

Stress response in Drosophila subobscura

The pattern of puffing and protein synthesis was determined in individuals of Drosophila subobscura subjected to heat shock. Depending on the extent of the heat treatment, the response at the puffing level varied. Some puffs were expressed at 31°–34°C, and others at 37° C. Considering the response as a whole the depression of gene activity after shock at 31°–34° C in individuals raised at 19° C was greater than with the other treatments. Six major heat shock proteins (Hsps) were found in this species. The properties of the high molecular weight proteins are conserved their electrophoretic characteristics and the range of temperatures over which they are synthesized are close to those in other Drosophila species. Remarkable heterogeneity was found in the small Hsps. In addition, an M_r=41000 Hsp was clearly identified in this species. A low level of variability in the patterns of protein synthesis compared with those of puffing activity was detected.     

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.     

Chromosomal inversion polymorphism of Drosophila subobscura from south‐eastern part of Europe

Five Drosophila subobscura populations from south‐eastern Europe were investigated with respect to chromosomal inversion polymorphism. They belong to central European populations and their gene arrangement frequencies are in accordance with latitudinal variation observed in this species. The new complex gene arrangement A₂₊₈₊₉, found in one of the populations analysed, may be specific for the southern Balkan refugium. A general tendency of gene arrangement frequencies to exhibit some southern characteristics was found for some of the autosomal standard arrangements.     

Global warming and chromosomal inversion adaptation in isolated islands: Drosophila subobscura populations from Madeira

Global warming is an environmental phenomenon to which species must adapt to survive. Drosophila subobscura presents an adaptive capacity due to its chromosomal inversion polymorphism. Until now, the impact of global warming on this polymorphism has been studied in D. subobscura populations located either on a continental mainland or on islands not far from a continent. In this context, gene flow could be a relevant mechanism allowing the movement of thermally adapted inversions between populations. Our aim was to sample and study the chromosomal polymorphism on Madeira, a small isolated island in the Atlantic Ocean. We compared our findings with those reported in the same location approximately four and five decades ago. Moreover, we studied whether global warming has occurred on this island by analyzing mean, maximum and minimum temperatures over a 55‐year period. All atmospheric parameters have increased significantly, consistent with climate change expectations. Frequencies and chromosomal thermal index values of thermal adapted inversions remained quite stable over years. Furthermore, J, U and O chromosomes are almost fixed for “warm” adapted inversions. Thus, if there is little genetic variability remaining and temperatures continue increasing, island populations of D. subobscura might be on the threshold of endangerment. However, apart from selection, genetic drift and inbreeding, other processes, such as phenotypic plasticity or thermoregulatory behavior, could be involved in the survival of the species’ populations. Finally, although in danger, D. subobscura is a generalist that lives in humanized environments, and this fact could favor its persistence on Madeira Island.     

The genetics of Drosophila subobscura populations XVIII. Multiple insemination and sperm displacement in Drosophila subobscura

The multiple insemination of Drosophila subobscura females in nature was studied by the analysis of single female offspring using two highly polymorphic enzyme systems. On qualitative grounds it was found that 23% of these females were inseminated at least twice. The power of detection of one enzyme system was estimated using the already known multi-inseminated females detected by the other system. This permitted the true frequency estimation of multi-inseminated females, in nature, which is 0.276 (for 95% confidence limits it ranges from 0.186 to 0.417). In a laboratory experiment it was found that sperm from the second inseminating male displaces the sperm from the first male.     

Population structure of Drosophila subobscura: Non-random microdispersion of inversion polymorphism on a mountain slope

On a forested mountain slope in the Swiss Alps gene arrangements of D. subobscura were sampled from altitudes between 600 and 1 900 m above sea level during the years 1977 and 1979. In both years the standard arrangement of chromosome I increased in frequency with altitude. Judging from the pattern of gene arrangement frequencies the population was spatially structured on a fine scale within altitude and on a coarser scale between altitudes. There were temporal changes between years. In spite of the fact that D. subobscura is at its ecological margin at the timberline no evidence of reduced variability was found there.     

Chromosomal inversion polymorphism of Drosophila subobscura populations from Jastrebac Mountain shows temporal and habitat-related changes

Inversion polymorphism in populations of D. subobscura from a beech forest on Jastrebac mountain was studied in June 1990, 1993, and 1994, respectively. The same analysis was performed in 1990 for D. subobscura populations in a beech forest and an oak forest in the same region. Statistically significant differences in the frequencies of the gene arrangements of A₁, J and U chromosome were observed during the period of investigation. A tendency towards a decrease in the frequency of the standard gene arrangements was found for all chromosomes, but was particularly evident with chromosomes A and J. The frequency of the gene arrangements A₁ A₂, J₁ and U_1–2+6 increased at the same time. Differences in the frequency of the gene arrangements of A, J and U chromosomes were also observed when the populations from two ecologically different habitats (beech and oak forest) were compared in 1990.     

Evolutionary history of the third chromosome gene arrangements of Drosophila pseudoobscura inferred from inversion breakpoints

The third chromosome of Drosophila pseudoobscura is polymorphic for numerous gene arrangements that form classical clines in North America. The polytene salivary chromosomes isolated from natural populations revealed changes in gene order that allowed the different gene arrangements to be linked together by paracentric inversions representing one of the first cases where genetic data were used to construct a phylogeny. Although the inversion phylogeny can be used to determine the relationships among the gene arrangements, the cytogenetic data are unable to infer the ancestral arrangement or the age of the different chromosome types. These are both important properties if one is to infer the evolutionary forces responsible for the spread and maintenance of the chromosomes. Here, we employ the nucleotide sequences of 18 regions distributed across the third chromosome in 80-100 D. pseudoobscura strains to test whether five gene arrangements are of unique or multiple origin, what the ancestral arrangement was, and what are the ages of the different arrangements. Each strain carried one of six commonly found gene arrangements and the sequences were used to infer their evolutionary relationships. Breakpoint regions in the center of the chromosome supported monophyly of the gene arrangements, whereas regions at the ends of the chromosome gave phylogenies that provided less support for monophyly of the chromosomes either because the individual markers did not have enough phylogenetically informative sites or genetic exchange scrambled information among the gene arrangements. A data set where the genetic markers were concatenated strongly supported a unique origin of the different gene arrangements. The inversion polymorphism of D. pseudoobscura is estimated to be about a million years old. We have also shown that the generated phylogeny is consistent with the cytological phylogeny of this species. In addition, the data presented here support hypothetical as the ancestral arrangement. One of the youngest arrangements, Arrowhead, has one of the highest population frequencies suggesting that selection has been responsible for its rapid increase.