Genetic basis and repeatability for desiccation resistance in Drosophila melanogaster (Diptera: Drosophilidae).

Dehydration is a stress factor for organisms inhabiting natural habitats where water is scarce. Thus, it may be expected that species facing arid environments will develop mechanisms that maximize resistance to desiccation. Insects are excellent models for studying the effects of dehydration as well as the mechanisms and processes that prevent water loss since the effect of desiccation is greater due to the higher area/volume ratio than larger animals. Even though physiological and behavioral mechanisms to cope with desiccation are being understood, the genetic basis underlying the mechanisms related to variation in desiccation resistance and the context-dependent effect remain unsolved. Here we analyze the genetic bases of desiccation resistance in Drosophila melanogaster and identify candidate genes that underlie trait variation. Our quantitative genetic analysis of desiccation resistance revealed sexual dimorphism and extensive genetic variation. The phenotype-genotype association analyses (GWAS) identified 71 candidate genes responsible for total phenotypic variation in desiccation resistance. Half of these candidate genes were sex-specific suggesting that the genetic architecture underlying this adaptive trait differs between males and females. Moreover, the public availability of desiccation data analyzed on the same lines but in a different lab allows us to investigate the reliability and repeatability of results obtained in independent screens. Our survey indicates a pervasive micro-environment lab-dependent effect since we did not detect overlap in the sets of genes affecting desiccation resistance identified between labs.

The relationship between morphology and flight in Drosophila: a study of two pairs of sibling species from a natural population.

Insect flight is a complex trait involved in different behaviors, from the search for sexual partners, food, or breeding sites. Many studies have postulated the adaptive advantages of certain morphological traits in relation to increased flight capacity, such as low values of wing loading or high values of wing:thorax ratio and wing-aspect ratio. However, few studies have evaluated the relationship between variables related to flight and morphological traits in Drosophila. This work aimed to study morphological traits in males and females of two pairs of sibling species: Drosophila buzzatii Patterson and Wheeler-Drosophila koeferae Fontdevila and Wasserman, and Drosophila melanogaster Meigen-Drosophila simulans Sturtevant, and to analyze its relationship with flight. We detected the highest proportion of flight time in D. koepferae and D. simulans compared to D. buzzatii and D. melanogaster, respectively. Our results also revealed sexual dimorphism, with males exhibiting a higher proportion of flight time than females. Surprisingly, we did not find a general pattern to explain the relationship between morphology and the proportion of flight time because associations varied depending upon the analyses (considering all groups together or each sex-species combination separately). Moreover, these associations explained a low percentage of variation, suggesting that other nonmorphological components related to flight, such as physiological variables, should be taken into account. This work allowed us to show the variability and complexity of an aspect of flight, suggesting that the adaptive role of the morphological traits studied might have been overestimated.

Contrasting levels of genotype by environment interaction for life history and morphological traits in invasive populations of Zaprionus indianus (Diptera: Drosophilidae).

It has been demonstrated that phenotypic plasticity and genotype by environment interaction are important for coping with new and heterogeneous environments during invasions. Zaprionus indianus Gupta (Diptera: Drosophilidae) is an Afrotropical invasive fly species introduced to the South American continent in 1999. This species is generalist and polyphagous, since it develops and feeds in several different fruit species. These characteristics of Z. indianus suggest that phenotypic plasticity and genotype by environment interaction may be important in this species invasion process. In this sense, our aim was to investigate the role of genetic variation for phenotypic plasticity (genotype by environment interaction) in Z. indianus invasion of the South American continent. Specifically, we quantified quantitative genetic variation and genotype by environment interactions of morphological and life history traits in different developmental environments, that is, host fruits. This was done in different populations in the invasive range of Z. indianus in Argentina. Results showed that Z. indianus populations have considerable amounts of quantitative genetic variation. Also, genotype by environment interactions was detected for the different traits analyzed in response to the different developmental environments. Interestingly, the amounts and patterns of these parameters differed between populations. We interpreted these results as the existence of differences in evolutionary potential between populations that have an important role in the short- and long-term success of the Z. indianus invasion process.

Stage- and thermal-specific genetic architecture for preadult viability in natural populations of Drosophila melanogaster.

Studying the processes affecting variation for preadult viability is essential to understand the evolutionary trajectories followed by natural populations. This task requires focusing on the complex nature of the phenotype-genotype relationship by taking into account usually neglected aspects of the phenotype and recognizing the modularity between different ontogenetic stages. Here, we describe phenotypic variability for viability during the larval and pupal stages in lines derived from three natural populations of Drosophila melanogaster, as well as the variability for phenotypic plasticity and canalization at two different rearing temperatures. The results indicate that the three populations present significant phenotypic differences for preadult viability. Furthermore, distinct aspects of the phenotype (means, plasticity, canalization, plasticity of canalization) are affected by different genetic bases underlying changes in viability in a stage- and environment-specific manner. These findings explain the generalized maintenance of genetic variability for this fitness trait.

Phenotypic plasticity in Drosophila cactophilic species: the effect of competition, density, and breeding sites.

Changes in the environmental conditions experienced by naturally occurring populations are frequently accompanied by changes in adaptive traits allowing the organism to cope with environmental unpredictability. Phenotypic plasticity is a major aspect of adaptation and it has been involved in population dynamics of interacting species. In this study, phenotypic plasticity (i.e., environmental sensitivity) of morphological adaptive traits were analyzed in the cactophilic species Drosophila buzzatii and Drosophila koepferae (Diptera: Drosophilidae) considering the effect of crowding conditions (low and high density), type of competition (intraspecific and interspecific competition) and cacti hosts (Opuntia and Columnar cacti). All traits (wing length, wing width, thorax length, wing loading and wing aspect) showed significant variation for each environmental factor considered in both Drosophila species. The phenotypic plasticity pattern observed for each trait was different within and between these cactophilic Drosophila species depending on the environmental factor analyzed suggesting that body size-related traits respond almost independently to environmental heterogeneity. The effects of ecological factors analyzed in this study are discussed in order to elucidate the causal factors investigated (type of competition, crowding conditions and alternative host) affecting the election of the breeding site and/or the range of distribution of these cactophilic species.

Natural Genetic Variation and Candidate Genes for Morphological Traits in Drosophila melanogaster.

Body size is a complex character associated to several fitness related traits that vary within and between species as a consequence of environmental and genetic factors. Latitudinal and altitudinal clines for different morphological traits have been described in several species of Drosophila and previous work identified genomic regions associated with such variation in D. melanogaster. However, the genetic factors that orchestrate morphological variation have been barely studied. Here, our main objective was to investigate genetic variation for different morphological traits associated to the second chromosome in natural populations of D. melanogaster along latitudinal and altitudinal gradients in Argentina. Our results revealed weak clinal signals and a strong population effect on morphological variation. Moreover, most pairwise comparisons between populations were significant. Our study also showed important within-population genetic variation, which must be associated to the second chromosome, as the lines are otherwise genetically identical. Next, we examined the contribution of different candidate genes to natural variation for these traits. We performed quantitative complementation tests using a battery of lines bearing mutated alleles at candidate genes located in the second chromosome and six second chromosome substitution lines derived from natural populations which exhibited divergent phenotypes. Results of complementation tests revealed that natural variation at all candidate genes studied, invected, Fasciclin 3, toucan, Reticulon-like1, jing and CG14478, affects the studied characters, suggesting that they are Quantitative Trait Genes for morphological traits. Finally, the phenotypic patterns observed suggest that different alleles of each gene might contribute to natural variation for morphological traits. However, non-additive effects cannot be ruled out, as wild-derived strains differ at myriads of second chromosome loci that may interact epistatically with mutant alleles.

Gene-by-temperature interactions and candidate plasticity genes for morphological traits in Drosophila melanogaster.

Understanding the genetic architecture of any quantitative trait requires identifying the genes involved in its expression in different environmental conditions. This goal can be achieved by mutagenesis screens in genetically tractable model organisms such as Drosophila melanogaster. Temperature during ontogenesis is an important environmental factor affecting development and phenotypic variation in holometabolous insects. In spite of the importance of phenotypic plasticity and genotype by environment interaction (GEI) for fitness related traits, its genetic basis has remained elusive. In this context, we analyzed five different adult morphological traits (face width, head width, thorax length, wing size and wing shape) in 42 co-isogenic single P-element insertional lines of Drosophila melanogaster raised at 17°C and 25°C. Our analyses showed that all lines differed from the control for at least one trait in males or females at either temperature. However, no line showed those differences for all traits in both sexes and temperatures simultaneously. In this sense, the most pleiotropic candidate genes were CG34460, Lsd-2 and Spn. Our analyses also revealed extensive genetic variation for all the characters mostly indicated by strong GEIs. Further, our results indicate that GEIs were predominantly explained by changes in ranking order in all cases suggesting that a moderate number of genes are involved in the expression of each character at both temperatures. Most lines displayed a plastic response for at least one trait in either sex. In this regard, P-element insertions affecting plasticity of a large number of traits were associated to the candidate genes Btk29A, CG43340, Drak and jim. Further studies will help to elucidate the relevance of these genes on the morphogenesis of different body structures in natural populations of D. melanogaster.

Genetic variation in heat-stress tolerance among South American Drosophila populations.

Spatial or temporal differences in environmental variables, such as temperature, are ubiquitous in nature and impose stress on organisms. This is especially true for organisms that are isothermal with the environment, such as insects. Understanding the means by which insects respond to temperature and how they will react to novel changes in environmental temperature is important for understanding the adaptive capacity of populations and to predict future trajectories of evolutionary change. The organismal response to heat has been identified as an important environmental variable for insects that can dramatically influence life history characters and geographic range. In the current study we surveyed the amount of variation in heat tolerance among Drosophila melanogaster populations collected at diverse sites along a latitudinal gradient in Argentina (24°-38°S). This is the first study to quantify heat tolerance in South American populations and our work demonstrates that most of the populations surveyed have abundant within-population phenotypic variation, while still exhibiting significant variation among populations. The one exception was the most heat tolerant population that comes from a climate exhibiting the warmest annual mean temperature. All together our results suggest there is abundant genetic variation for heat-tolerance phenotypes within and among natural populations of Drosophila and this variation has likely been shaped by environmental temperature.

Stage-specific effects of candidate heterochronic genes on variation in developmental time along an altitudinal cline of Drosophila melanogaster.

BACKGROUND: Previously, we have shown there is clinal variation for egg-to-adult developmental time along geographic gradients in Drosophila melanogaster. Further, we also have identified mutations in genes involved in metabolic and neurogenic pathways that affect development time (heterochronic genes). However, we do not know whether these loci affect variation in developmental time in natural populations. METHODOLOGY/PRINCIPAL FINDINGS: Here, we constructed second chromosome substitution lines from natural populations of Drosophila melanogaster from an altitudinal cline, and measured egg-adult development time for each line. We found not only a large amount of genetic variation for developmental time, but also positive associations of the development time with thermal amplitude and altitude. We performed genetic complementation tests using substitution lines with the longest and shortest developmental times and heterochronic mutations. We identified segregating variation for neurogenic and metabolic genes that largely affected the duration of the larval stages but had no impact on the timing of metamorphosis. CONCLUSIONS/SIGNIFICANCE: Altitudinal clinal variation in developmental time for natural chromosome substitution lines provides a unique opportunity to dissect the response of heterochronic genes to environmental gradients. Ontogenetic stage-specific variation in invected, mastermind, cricklet and CG14591 may affect natural variation in development time and thermal evolution.

Identifying candidate genes affecting developmental time in Drosophila melanogaster: pervasive pleiotropy and gene-by-environment interaction.

BACKGROUND: Understanding the genetic architecture of ecologically relevant adaptive traits requires the contribution of developmental and evolutionary biology. The time to reach the age of reproduction is a complex life history trait commonly known as developmental time. In particular, in holometabolous insects that occupy ephemeral habitats, like fruit flies, the impact of developmental time on fitness is further exaggerated. The present work is one of the first systematic studies of the genetic basis of developmental time, in which we also evaluate the impact of environmental variation on the expression of the trait. RESULTS: We analyzed 179 co-isogenic single P[GT1]-element insertion lines of Drosophila melanogaster to identify novel genes affecting developmental time in flies reared at 25 degrees C. Sixty percent of the lines showed a heterochronic phenotype, suggesting that a large number of genes affect this trait. Mutant lines for the genes Merlin and Karl showed the most extreme phenotypes exhibiting a developmental time reduction and increase, respectively, of over 2 days and 4 days relative to the control (a co-isogenic P-element insertion free line). In addition, a subset of 42 lines selected at random from the initial set of 179 lines was screened at 17 degrees C. Interestingly, the gene-by-environment interaction accounted for 52% of total phenotypic variance. Plastic reaction norms were found for a large number of developmental time candidate genes. CONCLUSION: We identified components of several integrated time-dependent pathways affecting egg-to-adult developmental time in Drosophila. At the same time, we also show that many heterochronic phenotypes may arise from changes in genes involved in several developmental mechanisms that do not explicitly control the timing of specific events. We also demonstrate that many developmental time genes have pleiotropic effects on several adult traits and that the action of most of them is sensitive to temperature during development. Taken together, our results stress the need to take into account the effect of environmental variation and the dynamics of gene interactions on the genetic architecture of this complex life-history trait.

Dynamic genetic interactions determine odor-guided behavior in Drosophila melanogaster.

Understanding the genetic architecture of complex traits requires identification of the underlying genes and characterization of gene-by-gene and genotype-by-environment interactions. Behaviors that mediate interactions between organisms and their environment are complex traits expected to be especially sensitive to environmental conditions. Previous studies on the olfactory avoidance response of Drosophila melanogaster showed that the genetic architecture of this model behavior depends on epistatic networks of pleiotropic genes. We performed a screen of 1339 co-isogenic p[GT1]-element insertion lines to identify novel genes that contribute to odor-guided behavior and identified 55 candidate genes with known p[GT1]-element insertion sites. Characterization of the expression profiles of 10 p[GT1]-element insertion lines showed that the effects of the transposon insertions are often dependent on developmental stage and that hypomorphic mutations in developmental genes can elicit profound adult behavioral deficits. We assessed epistasis among these genes by constructing all possible double heterozygotes and measuring avoidance responses under two stimulus conditions. We observed enhancer and suppressor effects among subsets of these P-element-tagged genes, and surprisingly, epistatic interactions shifted with changes in the concentration of the olfactory stimulus. Our results show that the manifestation of epistatic networks dynamically changes with alterations in the environment.

The genetic architecture of odor-guided behavior in Drosophila: epistasis and the transcriptome.

We combined transcriptional profiling and quantitative genetic analysis to elucidate the genetic architecture of olfactory behavior in Drosophila melanogaster. We applied whole-genome expression analysis to five coisogenic smell-impaired (smi) mutant lines and their control. We used analysis of variance to partition variation in transcript abundance between males and females and between smi genotypes and to determine the genotype-by-sex interaction. A total of 666 genes showed sexual dimorphism in transcript abundance, and 530 genes were coregulated in response to one or more smi mutations, showing considerable epistasis at the level of the transcriptome in response to single mutations. Quantitative complementation tests of mutations at these coregulated genes with the smi mutations showed that in most cases (67%) epistatic interactions for olfactory behavior mirrored epistasis at the level of transcription, thus identifying new candidate genes regulating olfactory behavior.

Vanaso is a candidate quantitative trait gene for Drosophila olfactory behavior.

Most animals depend on olfaction for survival and procreation. Odor-guided behavior is a quantitative trait, with phenotypic variation due to multiple segregating quantitative trait loci (QTL). Despite its profound biological importance, the genetic basis of naturally occurring variation in olfactory behavior remains unexplored. Here, we mapped a single Drosophila QTL affecting variation in avoidance response to benzaldehyde, using a population of recombinant inbred lines. Deficiency complementation mapping resolved this region into one female- and one male-specific QTL. Subsequent quantitative complementation tests to all available mutations of positional candidate genes showed that the female-specific QTL failed to complement a P-element insertional mutation, l(3)04276. The P-element insertion was in the intron of a novel gene, Vanaso, which contains a putative guanylate binding protein domain, is highly polymorphic, and is expressed in the third antennal segment, the major olfactory organ of Drosophila. No expression was detected in the fly brain, suggesting that Vanaso plays a role in peripheral chemosensory processes rather than in central integration of olfactory information. QTL mapping followed by quantitative complementation tests to deficiencies and mutations is an effective strategy for gene discovery that allows characterization of effects of recessive lethal genes on adult phenotypes and here enabled identification of a candidate gene that contributes to sex-specific quantitative variation in olfactory behavior.