A clinally varying promoter polymorphism associated with adaptive variation in wing size in Drosophila

Body size often shows adaptive clines in many ectotherms across altitude and latitude, but little is known about the genetic basis of these adaptive clines. Here we identify a polymorphism in the Dca (Drosophila cold acclimation) gene in Drosophila melanogaster that influences wing size, affects wing:thorax allometry and also controls a substantial proportion of the clinal wing‐size variation. A polymorphism in the promoter region of Dca had two common alleles showing strong reciprocal clinal variation in frequency with latitude along the east coast of Australia. The Dca‐237 allele increased towards the tropics where wing size is smaller. A within‐population association study highlighted that an increase in the frequency of this allele decreased wing size but did not influence thorax size. A manipulated increase in the level of expression of Dca achieved through UAS‐GAL4 was associated with a decrease in wing size but had no effect on thorax size. This was consistent with higher Dca expression levels in family lines with higher frequency of the Dca‐237 allele. Genetic variation in the promoter region of the Dca gene appears to influence adaptive size variation in the eastern Australian cline of Drosophila melanogaster and accounts for more than 10% of the genetic variation in size within and between populations.     

A comparative study of the short term cold resistance response in distantly related Drosophila species: the role of regucalcin and frost

The molecular basis of short term cold resistance (indexed as chill-coma recovery time) has been mostly addressed in D. melanogaster, where candidate genes (Dca (also known as smp-30) and Frost (Fst)) have been identified. Nevertheless, in Drosophila, the ability to tolerate short term exposure to low temperatures evolved several times independently. Therefore, it is unclear whether variation in the same candidate genes is also responsible for short term cold resistance in distantly related Drosophila species. It should be noted that Dca is a candidate gene for cold resistance in the Sophophora subgenus only, since there is no orthologous gene copy in the Drosophila subgenus. Here we show that, in D. americana (Drosophila subgenus), there is a north-south gradient for a variant at the 5' non-coding region of regucalcin (a Dca-like gene; in D. melanogaster the proteins encoded by the two genes share 71.9% amino acid identities) but in our D. americana F2 association experiment there is no association between this polymorphism and chill-coma recovery times. Moreover, we found no convincing evidence that this gene is up-regulated after cold shock in both D. americana and D. melanogaster. Size variation in the Fst PEST domain (putatively involved in rapid protein degradation) is observed when comparing distantly related Drosophila species, and is associated with short term cold resistance differences in D. americana. Nevertheless, this effect is likely through body size variation. Moreover, we show that, even at two hours after cold shock, when up-regulation of this gene is maximal in D. melanogaster (about 48 fold expression change), in D. americana this gene is only moderately up-regulated (about 3 fold expression change). Our work thus shows that there are important differences regarding the molecular basis of cold resistance in distantly related Drosophila species.     

Candidate genes and thermal phenotypes: identifying ecologically important genetic variation for thermotolerance in the Australian Drosophila melanogaster cline

Clinal variation in traits often reflects climatic adaptation; in Drosophila melanogaster clinal variation provides an opportunity to link variation in chromosomal inversions, microsatellite loci and various candidate genes to adaptive variation in traits. We undertook association studies with crosses from a single population of D. melanogaster from eastern Australia to investigate the association between genetic markers and traits showing clinal variation. By genotyping parents and phenotyping offspring, we minimized genotyping costs but had the power to detect association between markers and quantitative traits. Consistent with prior studies, we found strong associations between the clinal chromosomal inversion In(3R)Payne and markers within it, as well as among these markers. We also found an association between In(3L)Payne and one marker located within this inversion. Of the five predicted associations between markers and traits, four were detected (increased heat, decreased cold resistance and body size with the heat shock gene hsr-omega S, increased cold resistance with the inversion In(3L)Payne), while one was not detected (heat resistance and the heat shock gene hsp68). In a set of eight exploratory tests, we detected one positive association (between hsp23a and heat resistance) but no associations of heat resistance with alleles at the hsp26, hsp83, Desat 2, alpha-Gpdh, hsp70 loci, while cold resistance was not associated with Frost and Dca loci. These results confirm interactions between hsr-omega and thermal resistance, as well as between In(3L)Payne and cold resistance, but do not provide evidence for associations between thermal responses and alleles at other clinically varying marker genes.     

应用果蝇yellow基因的分子模式分析基因转应作用(英文)

基因转应作用（ｔｒａｎｓｖｅｃｔｉｏｎ）是基因表达的一种方式,这种方式是由等位基因配对及其相互作用所介导的。基因转应作用的现象已在果蝇的多种基因中发现。这种作用可产生正负两种效应。而且,在其它物种中,也逐渐发现了类似的现象。例如,在植物中的基因沉默现象（ｇｅｎｅｓｉｌｅｎｃｉｎｇ）以及在小鼠中的基因转激活作用（ｔｒａｎｓａｃｔｉｖａｔｉｏｎ）等。因此,阐明基因转应作用的机理,将有助于了解基因表达调节及增强子调控活动的分子基础。本文应用果蝇ｙｅｌｏｗ基因为模式来探讨基因转应作用的分子机制。前期研究表明,ｙｅｌｏｗ基因转应作用发生于ｇｙｐｓｙ诱导的ｙ２突变种和ｙｅｌｏｗ亚等位基因（ｙｈ）之间。为了证实是否ｇｙｐｓｙ是基因转应作用所必需的ＤＮＡ元件,我们鉴定了一种新的ｙｅｌｏｗ突变种,称为ｙ２３７４。ｙ２３７４突变是一种基因表达的组织特异性改变,这一改变使ｙ２３７４果蝇在翅和身体部位表皮着色呈突变型。通过遗传分析表明,ｙ２３７４也可与ｙｈ（如ｙ１＃８）产生基因转应作用。ｙ１＃８是一种无效的ｙｅｌｏｗ等位基因,它包含一个启动子和部分编码区序列的缺失。然而,当ｙ２３７４与ｙ１＃８杂交后,其杂交后代的表现型可由ｙ２３７     

The impact of shared ancestral variation on hybrid male lethality--a 16 codon indel in the Drosophila simulans Lhr gene

The Lethal hybrid rescue (Lhr) gene causes hybrid male lethality in crosses between Drosophila simulans and D. melanogaster. Lhr(2) is a D. simulans allele, which rescues hybrid males. It has been recently proposed that a 16 codon insertion, which distinguishes the D. melanogaster and the canonical D. simulans allele, and is lacking in Lhr(2), may be responsible for the functional divergence of D. melanogaster and D. simulans Lhr alleles. Here, we show that the Lhr(2) allele lacking the insertion represents an ancestral polymorphism segregating at a moderate frequency in D. simulans. Crosses of D. melanogaster females to males from two D. simulans strains carrying this deletion showed a severe deficiency of viable hybrid males. Our results suggest that the absence of this insertion alone is not sufficient to explain functional differences between D. melanogaster and D. simulans Lhr alleles.     

Association between nucleotide variation in Egfr and wing shape in Drosophila melanogaster

As part of an effort to dissect quantitative trait locus effects to the nucleotide level, association was assessed between 238 single-nucleotide and 20 indel polymorphisms spread over 11 kb of the Drosophila melanogaster Egfr locus and nine relative warp measures of wing shape. One SNP in a conserved potential regulatory site for a GAGA factor in the promoter of alternate first exon 2 approaches conservative experiment-wise significance (P < 0.00003) in the sample of 207 lines for association with the location of the crossveins in the central region of the wing. Several other sites indicate marginal association with one or more other aspects of shape. No strong effects of sex or population of origin were detected with measures of shape, but two different sites were strongly associated with overall wing size in interaction with these fixed factors. Whole-gene sequencing in very large samples, rather than selective genotyping, would appear to be the only strategy likely to be successful for detecting subtle associations in species with high polymorphism and little haplotype structure. However, these features severely limit the ability of linkage disequilibrium mapping in Drosophila to resolve quantitative effects to single nucleotides.     

Regulation of genome stability by TEL1 and MEC1, yeast homologs of the mammalian ATM and ATR genes

DNA sequence surveys of Drosophila melanogaster populations show a strong positive correlation between the recombination rate experienced by a locus and its level of nucleotide polymorphism. In particular, surveys of the fourth chromosome gene ci(D) show greatly reduced levels of nucleotide variation; this observation was originally interpreted in terms of selective sweeps occurring on the nonrecombining fourth chromosome. Subsequent theoretical work has, however, uncovered several other selective processes that can reduce variation. In this study, we revisit the Drosophila fourth chromosome, investigating variation in 5-6 kb of the gene ankyrin in D. melanogaster and D. simulans. Silent nucleotide site diversity is approximately 5 x 10(-4) for both species, consistent with the previous observations of low variation at ci(D). Given the observed frequency spectra at ankyrin, coalescent simulations indicate that reduced diversity in the region is unlikely to be due to a selective sweep alone. We find evidence for recombinational exchange at this locus, and both species appear to be fixed for an insertion of the transposable element HB in an intron of ankyrin.     

DNA sequence variation and latitudinal associations in hsp23, hsp26 and hsp27 from natural populations of Drosophila melanogaster

Heat shock genes are considered to be likely candidate genes for environmental stress resistance. Nucleotide variation in the coding sequence of the small heat shock genes (hsps) hsp26 and hsp27 from Drosophila melanogaster was studied in flies originating from the Netherlands and eastern Australia. The hsp26 gene was polymorphic for an insertion/deletion of three extra amino acids and two nonsynonymous changes in all populations. The hsp27 gene exhibited two nonsynonymous changes and three synonymous mutations. The hsp26 polymorphism showed a latitudinal cline along the east coast of Australia. This pattern was not confounded by the fact that the shsps are located in the inversion In(3 L)P which also shows a latitudinal cline in eastern Australia. A similar latitudinal cline was found for the previously described variation in hsp23, while frequencies of hsp27 alleles did not change with latitude. These findings suggest that variation at two of the shsps or closely linked loci are under selection in natural populations of D. melanogaster.     

In search of clinal variation in the period and clock timing genes in Australian Drosophila melanogaster populations

Clinal variation for repeat number in the Thr-Gly region of the period circadian timing gene in Drosophila melanogaster was described in Europe and has subsequently been used as evidence of thermal selection on period alleles. To test for clinal variation in this gene along the east coast of Australia, the period polymorphism was scored on flies from multiple samples collected repeatedly over a 5-year interval, along with variation at another circadian rhythm locus, clock. For period, there was no consistent evidence of clinal variation in the 17 and/or 20 repeat alleles, although when average allele length was examined a weak consistent clinal pattern was detected. For clock there was no evidence of clinal variation in the two most common alleles or in average repeat size. These data are inconsistent with the reported patterns in Europe and suggest that clinal variation in timing genes needs to be re-examined in this region.     

Replication of an Egfr-wing shape association in a wild-caught cohort of Drosophila melanogaster

Linkage disequilibrium mapping has been used extensively in medical and evolutionary genetics to map causal polymorphisms within genes associated with disease status or phenotypic variation for a trait. However, the initial findings of most nonhuman studies have not been replicated in subsequent studies, due in part to false positives, as well as additional factors that can render true positives unreplicable. These factors may be more severe when the initial study is performed using an experimental population of organisms reared under controlled lab conditions. We demonstrate that despite considerable phenotypic differences for wing shape between a lab-reared experimental population and a wild-caught cohort of Drosophila melanogaster, an association between a putative regulatory polymorphism in Egfr and wing shape can be replicated. These results are discussed both within the framework of future association-mapping studies and within the context of the evolutionary dynamics of alleles in populations.     

The Rosy Region of Drosophila Melanogaster and Drosophila Simulans. I. Contrasting Levels of Naturally Occurring DNA Restriction Map Variation and Divergence

A 40-kb region around the rosy and snake loci was analyzed for restriction map variation among 60 lines of Drosophila melanogaster and 30 lines of Drosophila simulans collected together at a single locality in Raleigh, North Carolina. DNA sequence variation in D. simulans was estimated to be 6.3 times greater than in D. melanogaster (heterozygosities per nucleotide of 1.9% vs. 0.3%). This result stands in marked contrast to results of studies of phenotypic variation including proteins (allozymes), morphology and chromosome arrangements which are generally less variable and less geographically differentiated in D. simulans. Intraspecific polymorphism is not distributed uniformly over the 40-kb region. The level of heterozygosity per nucleotide varies more than 12-fold across the region in D. simulans, being highest over the hsc2 gene. Similar, though less extreme, variation in heterozygosity is also observed in D. melanogaster. Average interspecific divergence (corrected for intraspecific polymorphism) averaged 3.8%. The pattern of interspecific divergence over the 40-kb region shows some disparities with the spatial distribution of intraspecific variation, but is generally consistent with selective neutrality predictions: the most polymorphic regions within species are generally the most divergent between species. Sequence-length polymorphism is observed for D. melanogaster to be at levels comparable to other gene regions in this species. In contrast, no sequence length variation was observed among D. simulans chromosomes (limit of resolution approximately 100 bp). These data indicate that transposable elements play at best a minor role in the generation of naturally occurring genetic variation in D. simulans compared to D. melanogaster. We hypothesize that differences in species effective population size are the major determinant of the contrasting levels and patterns of DNA sequence and insertion/deletion variation that we report here and the patterns of allozyme and morphological variation and differentiation reported by other workers for these two species.     

Temporal dynamics and variation of multilocus ISSR-PCR DNA markers in the Uman' population of Drosophila melanogaster over two decades (1984-2004)

The temporal dynamics of genomic variation in the Uman' (Ukraine) population of Drosophila melanogaster over the period 1984-2004 was studied using multilocus ISSR markers. Considerable polymorphism of the genomic DNA fragments corresponding to ISSR markers was found in the D. melanogaster population studied: the values of average heterozygosity varied from 0.085 to 0.127 depending on the year. Significant differences in the frequencies of dominant alleles between the samples of different years were recorded for 12 of the 30 DNA fractions detected. These changes are nondirectional and random. The pattern of detected variation suggests the determining influence of gene drift and migration process on the variation of noncoding DNA sequences in the Uman' population of D. melanogaster.     

An interspecific QTL study of Drosophila wing size and shape variation to investigate the genetic basis of morphological differences

The Drosophila wing has been used as a model in studies of morphogenesis and evolution; the use of such models can contribute to our understanding of mechanisms that promote morphological divergence among populations and species. We mapped quantitative trait loci (QTL) affecting wing size and shape traits using highly inbred introgression lines between D. simulans and D. sechellia, two sibling species of the melanogaster subgroup. Eighteen QTL peaks that are associated with 12 wing traits were identified, including two principal components. The wings of D. simulans and D. sechellia significantly diverged in size; two of the QTL peaks could account for part of this interspecific divergence. Both of these putative QTLs were mapped at the same cytological regions as other QTLs for intraspecific wing size variation identified in D. melanogaster studies. In these regions, one or more loci could account for intra- and interspecific variation in the size of Drosophila wings. Three other QTL peaks were related to a pattern of interspecific variation in wing size and shape traits that is summarized by one principal component. In addition, we observed that female wings are significantly larger and longer than male wings and the second, fourth and fifth longitudinal veins are closer together at the distal wing area. This pattern was summarized by another principal component, for which one QTL was mapped.     

QTL mapping reveals a striking coincidence in the positions of genomic regions associated with adaptive variation in body size in parallel clines of Drosophila melanogaster on different continents

Latitudinal genetic clines in body size are common in many ectotherm species and are attributed to climatic adaptation. Here, we use Quantitative Trait Loci (QTL) mapping to identify genomic regions associated with adaptive variation in body size in natural populations of Drosophila melanogaster from extreme ends of a cline in South America. Our results show that there is a significant association between the positions of QTL with strong effects on wing area in South America and those previously reported in a QTL mapping study of Australian cline end populations (P < 0.05). In both continents, the right arm of the third chromosome is associated with QTL with the strongest effect on wing area. We also show that QTL peaks for wing area and thorax length are associated with the same genomic regions, indicating that the clinal variation in the body size traits may have a similar genetic basis. The consistency of the results found for the South American and Australian cline end populations indicate that the genetic basis of the two clines may be similar and future efforts to identify the genes producing the response to selection should be focused on the genomic regions highlighted by the present work.     

Evolution of a desaturase involved in female pheromonal cuticular hydrocarbon biosynthesis and courtship behavior in Drosophila

Drosophila species exhibit polymorphism in female pheromonal cuticular hydrocarbons, with 7-monoenes produced in Drosophila simulans and 7,11-dienes in most populations of Drosophila melanogaster (5,9-dienes in several African populations). A female-biased desaturase, desatF, expressed only in D. melanogaster is involved in the synthesis of 7,11-dienes. We investigated the role of desatF in 5,9-diene flies. We constructed a 5,9-diene strain knock-down for desatF and showed that desatF is involved in 5,9-diene formation. We also studied D. melanogaster/D. simulans hybrids. These hybrid females produced dienes and received normal courtship from D. melanogaster males, but copulation success was reduced. With D. simulans males, courtship was decreased and no copulation occurred. Hybrids with a chromosomal deletion of the D. melanogaster desatF gene had no dienes and received normal courtship from D. simulans males; depending on the D. simulans parental strain, 7-19% of them succeeded in mating. D. simulans desatF promoter region shows 21-23% gaps and 86-89% identity with D. melanogaster promoter region, the coding region 93-94% identity, depending on the strain. These differences could explain the functional polymorphism of desatF observed between both species, contributing to different cuticular hydrocarbon profiles, that constitute an effective barrier between species.     

Linkage disequilibrium patterns across a recombination gradient in African Drosophila melanogaster

Previous multilocus surveys of nucleotide polymorphism have documented a genome-wide excess of intralocus linkage disequilibrium (LD) in Drosophila melanogaster and D. simulans relative to expectations based on estimated mutation and recombination rates and observed levels of diversity. These studies examined patterns of variation from predominantly non-African populations that are thought to have recently expanded their ranges from central Africa. Here, we analyze polymorphism data from a Zimbabwean population of D. melanogaster, which is likely to be closer to the standard population model assumptions of a large population with constant size. Unlike previous studies, we find that levels of LD are roughly compatible with expectations based on estimated rates of crossing over. Further, a detailed examination of genes in different recombination environments suggests that markers near the telomere of the X chromosome show considerably less linkage disequilibrium than predicted by rates of crossing over, suggesting appreciable levels of exchange due to gene conversion. Assuming that these populations are near mutation-drift equilibrium, our results are most consistent with a model that posits heterogeneity in levels of exchange due to gene conversion across the X chromosome, with gene conversion being a minor determinant of LD levels in regions of high crossing over. Alternatively, if levels of exchange due to gene conversion are not negligible in regions of high crossing over, our results suggest a marked departure from mutation-drift equilibrium (i.e., toward an excess of LD) in this Zimbabwean population. Our results also have implications for the dynamics of weakly selected mutations in regions of reduced crossing over.