Deficiency mapping of the genomic regions associated with effects on developmental stability in Drosophila melanogaster

Developmental stability is the tendency of morphological traits to resist the effects of developmental noise, and is commonly evaluated by examining fluctuating asymmetry (FA)-random deviations from perfect bilateral symmetry. Molecular mechanisms that control FA have been a long-standing topic of debate in the field of evolutionary biology and quantitative genetics. In this study, we mapped genomic regions associated with effects on the mean and FA of morphological traits, and characterized the trait specificity of those regions. A collection of isogenic deficiency strains established by the DrosDel project was used for deficiency mapping of genome regions associated with effects on FA. We screened 435 genome deficiencies or approximately 64.9% of the entire genome of Drosophila melanogaster to map the region that demonstrated a significant effect on FA of morphological traits. We found that 406 deficiencies significantly affected the mean of morphological traits, and 92 deficiencies increased FA. These results suggest that several genomic regions have the potential to affect developmental stability. They also suggest the possibility of the existence of trait-specific and trait-nonspecific mechanisms for stabilizing developmental processes. The new findings in this study could provide insight into the understanding of the genetic architecture underlying developmental stability.     

Novel genetic capacitors and potentiators for the natural genetic variation of sensory bristles and their trait specificity in Drosophila melanogaster

Cryptic genetic variation (CGV) is defined as the genetic variation that has little effect on phenotypic variation under a normal condition, but contributes to heritable variation under environmental or genetic perturbations. Genetic buffering systems that suppress the expression of CGV and store it in a population are called genetic capacitors, and the opposite systems are called genetic potentiators. One of the best‐known candidates for a genetic capacitor and potentiator is the molecular chaperone protein, HSP90, and one of its characteristics is that it affects the genetic variation in various morphological traits. However, it remains unclear whether the wide‐ranging effects of HSP90 on a broad range of traits are a general feature of genetic capacitors and potentiators. In the current study, I searched for novel genetic capacitors and potentiators for quantitative bristle traits of Drosophila melanogaster and then investigated the trait specificity of their genetic buffering effect. Three bristle traits of D. melanogaster were used as the target traits, and the genomic regions with genetic buffering effects were screened using the 61 genomic deficiencies examined previously for genetic buffering effects in wing shape. As a result, four and six deficiencies with significant effects on increasing and decreasing the broad‐sense heritability of the bristle traits were identified, respectively. Of the 18 deficiencies with significant effects detected in the current study and/or by the previous study, 14 showed trait‐specific effects, and four affected the genetic buffering of both bristle traits and wing shape. This suggests that most genetic capacitors and potentiators exert trait‐specific effects, but that general capacitors and potentiators with effects on multiple traits also exist.     

Multiple capacitors for natural genetic variation in Drosophila melanogaster

Cryptic genetic variation (CGV) or a standing genetic variation that is not ordinarily expressed as a phenotype is released when the robustness of organisms is impaired under environmental or genetic perturbations. Evolutionary capacitors modulate the amount of genetic variation exposed to natural selection and hidden cryptically; they have a fundamental effect on the evolvability of traits on evolutionary timescales. In this study, I have demonstrated the effects of multiple genomic regions of Drosophila melanogaster on CGV in wing shape. I examined the effects of 61 genomic deficiencies on quantitative and qualitative natural genetic variation in the wing shape of D. melanogaster. I have identified 10 genomic deficiencies that do not encompass a known candidate evolutionary capacitor, Hsp90, exposing natural CGV differently depending on the location of the deficiencies in the genome. Furthermore, five genomic deficiencies uncovered qualitative CGV in wing morphology. These findings suggest that CGV in wing shape of wild‐type D. melanogaster is regulated by multiple capacitors with divergent functions. Future analysis of genes encompassed by these genomic regions would help elucidate novel capacitor genes and better understand the general features of capacitors regarding natural genetic variation.     

Genome-wide deficiency mapping of the regions responsible for temporal canalization of the developmental processes of Drosophila melanogaster

Developmental processes of organisms are programmed to proceed in a finely regulated manner and finish within a certain period of time depending on the ambient environmental conditions. Therefore, variation in the developmental period under controlled genetic and environmental conditions indicates innate instability of the developmental process. In this study, we aimed to determine whether a molecular machinery exists that regulates the canalization of the developmental period and, if so, to test whether the same mechanism also stabilizes a morphological trait. To search for regions that influence the instability of the developmental period, we conducted genome-wide deficiency mapping with 441 isogenic deficiency strains covering 65.5% of the Drosophila melanogaster genome. We found that 11 independent deficiencies significantly increased the instability of the developmental period and 5 of these also significantly increased the fluctuating asymmetry of wing shape although there was no significant correlation between the instabilities of developmental period and wing shape in general. These results suggest that canalization processes of the developmental period and morphological traits are at least partially independent. Our findings emphasize the potential importance of temporal variation in development as an indicator of developmental stability and canalization and provide a novel perspective for understanding the regulation of phenotypic variability.     

Genetic mapping of adaptive wing size variation in Drosophila simulans

Many ecologically important traits exhibit latitudinal variation. Body size clines have been described repeatedly in insects across multiple continents, suggesting that similar selective forces are shaping these geographical gradients. It is unknown whether these parallel clinal patterns are controlled by the same or different genetic mechanism(s). We present here, quantitative trait loci (QTL) analysis of wing size variation in Drosophila simulans. Our results show that much of the wing size variation is controlled by a QTL on Chr 3L with relatively minor contribution from other chromosome arms. Comparative analysis of the genomic positions of the QTL indicates that the major QTL on Chr 3 are distinct in D. simulans and D. melanogaster, whereas the QTL on Chr 2R might overlap between species. Our results suggest that parallel evolution of wing size clines could be driven by non-identical genetic mechanisms but in both cases involve a major QTL as well as smaller effects of other genomic regions.     

Segregating variation in the polycomb group gene cramped alters the effect of temperature on multiple traits

The phenotype produced by a given genotype can be strongly modulated by environmental conditions. Therefore, natural populations continuously adapt to environment heterogeneity to maintain optimal phenotypes. It generates a high genetic variation in environment-sensitive gene networks, which is thought to facilitate evolution. Here we analyze the chromatin regulator crm, identified as a candidate for adaptation of Drosophila melanogaster to northern latitudes. We show that crm contributes to environmental canalization. In particular, crm modulates the effect of temperature on a genomic region encoding Hedgehog and Wingless signaling effectors. crm affects this region through both constitutive heterochromatin and Polycomb silencing. Furthermore, we show that crm European and African natural variants shift the reaction norms of plastic traits. Interestingly, traits modulated by crm natural variants can differ markedly between Drosophila species, suggesting that temperature adaptation facilitates their evolution.     

Countergradient variation in body shape between two populations of Atlantic cod (Gadus morhua)

Variation in morphological traits is generally thought to be cogradient, with environmental effects on phenotypic expression reinforcing genetic differences between populations. We compared body shape between two populations of Atlantic cod (Gadus morhua). Striking shape differences occurred between juveniles from the two populations when reared in a common laboratory environment. However, no difference in body shape occurred between wild-reared juveniles from the two populations, suggesting that the genetic differences between populations were obscured by opposing effects of the environmental differences experienced in the wild. We suggest that much of the genetic diversity in body shape of fishes may be cryptic, with stabilizing selection for the same optimal phenotype resulting in genetic divergence between populations subject to contrasting environmental influences.     

Extent of QTL Reuse During Repeated Phenotypic Divergence of Sympatric Threespine Stickleback

How predictable is the genetic basis of phenotypic adaptation? Answering this question begins by estimating the repeatability of adaptation at the genetic level. Here, we provide a comprehensive estimate of the repeatability of the genetic basis of adaptive phenotypic evolution in a natural system. We used quantitative trait locus (QTL) mapping to discover genomic regions controlling a large number of morphological traits that have diverged in parallel between pairs of threespine stickleback (Gasterosteus aculeatus species complex) in Paxton and Priest lakes, British Columbia. We found that nearly half of QTL affected the same traits in the same direction in both species pairs. Another 40% influenced a parallel phenotypic trait in one lake but not the other. The remaining 10% of QTL had phenotypic effects in opposite directions in the two species pairs. Similarity in the proportional contributions of all QTL to parallel trait differences was about 0.4. Surprisingly, QTL reuse was unrelated to phenotypic effect size. Our results indicate that repeated use of the same genomic regions is a pervasive feature of parallel phenotypic adaptation, at least in sticklebacks. Identifying the causes of this pattern would aid prediction of the genetic basis of phenotypic evolution.     

MicroRNA transgene overexpression complements deficiency-based modifier screens in Drosophila

Dosage-sensitive modifier screening is a powerful tool for linking genes to biological processes. Use of chromosomal deletions permits sampling the effects of removing groups of genes related by position on the chromosome. Here, we explore the use of inducible microRNA transgenes as a complement to deficiency-based modifier screens. miRNAs are predicted to have hundreds of targets. miRNA overexpression provides an efficient means to reduces expression of large gene sets. A collection of transgenes was prepared to allow overexpression of 89 miRNAs or miRNA clusters. These transgenes and a set of genomic deficiencies were screened for their ability to modify the bristle phenotype of the cell-cycle regulator minus. Sixteen miRNAs were identified as dominant suppressors, while the deficiency screen uncovered four genomic regions that contain a dominant suppressor. Comparing the genes uncovered by the deletions with predicted miRNA targets uncovered a small set of candidate suppressors. Two candidates were identified as suppressors of the minus phenotype, Cullin-4 and CG5199/Cut8. Additionally, we show that Cullin-4 acts through its substrate receptor Cdt2 to suppress the minus phenotype. We suggest that inducible microRNA transgenes are a useful complement to deficiency-based modifier screens.     

Evolution and maintenance of the environmental component of the phenotypic variance: benefit of plastic traits under changing environments

How environmental variances in quantitative traits are influenced by variable environments is an important problem in evolutionary biology. In this study, the evolution and maintenance of phenotypic variance in a plastic trait under stabilizing selection are investigated. The mapping from genotypic value to phenotypic value of the quantitative trait is approximated by a linear reaction norm, with genotypic effects on its phenotypic mean and sensitivity to environment. The environmental deviation is assumed to be decomposed into environmental quality, which interacts with genotypic value, and residual developmental noise, which is independent of genotype. Environmental quality and the optimal phenotype of stabilizing selection are allowed to randomly fluctuate in both space and time, and individuals migrate equally before development and reproduction among different niches. Analyses show that phenotypic plasticity is adaptive within variable environments if correlations have become established between the optimal phenotype and environmental quality in space and/or time. The evolved plasticity increases with variances in optimal phenotypes and correlations between optimal phenotype and environmental quality; this further induces increases in mean fitness and the environmental variance in the trait. Under certain circumstances, however, the environmental variance may decrease with increase in variation in environmental quality.     

Quantitative genetics of larval life-history traits in Rana temporaria in different environmental conditions

The degree to which genetic variation in a given trait varies among different populations of the same species and across different environments has seldom been quantified in wild vertebrate species. We investigated the expression of genetic variability and maternal effects in three larval life-history traits of the amphibian Rana temporaria. In a factorial laboratory experiment, five widely separated populations (max. 1600 km) were subjected to two different environmental treatments. Animal model analyses revealed that all traits were heritable (h(2) approximately 0.20) in all populations and under most treatment combinations. Although the cross-food treatment genetic correlations were close to unity, heritabilities under a restricted food regime tended to be lower than those under an ad libitum food regime. Likewise, maternal effects (m(2) approximately 0.05) were detected in most traits, and they tended to be most pronounced under restricted food conditions. We detected several cross-temperature genetic and maternal effects correlations that were lower than unity, suggesting that genotype-environment interactions and maternal effect-environment interactions are a significant source of phenotypic variation. The results reinforce the perspective that although the expression of genetic and maternal effects may be relatively homogeneous across different populations of the same species, local variation in environmental conditions can lead to significant variation in phenotypic expression of quantitative traits through genotype-environment and maternal effect-environment interactions.     

Apparent Genetic Complexity Generated by Developmental Thresholds: The Apterous Locus in DROSOPHILA MELANOGASTER

Mutations at the apterous (ap) locus in Drosophila melanogaster give rise to three distinct phenotypes: aberrant wings, female sterility and precocious adult death. The wing phenotype includes five types of abnormality: blistering, deficiencies, duplications, high-order repetitions and transformation of structures. The mildest phenotype is seen with homozygous apblt animals which have either normal or slightly blistered wings. Most alleles produce, in the homozygote, a deficient wing in which part or all of the wing margin and wing blade is missing, but wing hinge and notum regions are normal. Animals hemizygous for each of 20 ap alleles, as well as apID/apXa heterozygotes, show duplication of parts of the notum associated with complete wing deficiency. Animals heterozygous for apc and the other tested ap alleles show repetitions of parts of the anterior wing margin, an engrailed-like transformation of posterior wing margin into anterior margin or both. Both apblt and apc show similar phenotypes in homozygotes and hemizygotes, yet both produce a less extreme phenotype than that of the other hemizygotes, suggesting that neither mutation causes loss of the entire ap+ function. The 15 alleles that cause precocious death and female sterility occur in six complementation groups based on complementation for these phenotypes. This supports the previous conclusion that the effects of apterous mutations on the wing do not correlate with their effects on viability and fertility. We propose an explanation for the effects of apterous mutations on the wing in which quantitative reductions in the activity of gene product give rise to qualitatively different phenotypes because of different threshold requirements of the ap+ function for critical events in wing disc development.     

Genetic analysis of sunflower domestication

Quantitative trait loci (QTL) controlling phenotypic differences between cultivated sunflower and its wild progenitor were investigated in an F(3) mapping population. Composite interval mapping revealed the presence of 78 QTL affecting the 18 quantitative traits of interest, with 2-10 QTL per trait. Each QTL explained 3.0-68.0% of the phenotypic variance, although only 4 (corresponding to 3 of 18 traits) had effects >25%. Overall, 51 of the 78 QTL produced phenotypic effects in the expected direction, and for 13 of 18 traits the majority of QTL had the expected effect. Despite being distributed across 15 of the 17 linkage groups, there was a substantial amount of clustering among QTL controlling different traits. In several cases, regions influencing multiple traits harbored QTL with antagonistic effects, producing a cultivar-like phenotype for some traits and a wild-like phenotype for others. On the basis of the directionality of QTL, strong directional selection for increased achene size appears to have played a central role in sunflower domestication. None of the other traits show similar evidence of selection. The occurrence of numerous wild alleles with cultivar-like effects, combined with the lack of major QTL, suggests that sunflower was readily domesticated.     

Dissecting the Phenotypic Components of Crop Plant Growth and Drought Responses Based on High-Throughput Image Analysis

Significantly improved crop varieties are urgently needed to feed the rapidly growing human population under changing climates. While genome sequence information and excellent genomic tools are in place for major crop species, the systematic quantification of phenotypic traits or components thereof in a high-throughput fashion remains an enormous challenge. In order to help bridge the genotype to phenotype gap, we developed a comprehensive framework for high-throughput phenotype data analysis in plants, which enables the extraction of an extensive list of phenotypic traits from nondestructive plant imaging over time. As a proof of concept, we investigated the phenotypic components of the drought responses of 18 different barley (Hordeum vulgare) cultivars during vegetative growth. We analyzed dynamic properties of trait expression over growth time based on 54 representative phenotypic features. The data are highly valuable to understand plant development and to further quantify growth and crop performance features. We tested various growth models to predict plant biomass accumulation and identified several relevant parameters that support biological interpretation of plant growth and stress tolerance. These image-based traits and model-derived parameters are promising for subsequent genetic mapping to uncover the genetic basis of complex agronomic traits. Taken together, we anticipate that the analytical framework and analysis results presented here will be useful to advance our views of phenotypic trait components underlying plant development and their responses to environmental cues.     

Genomic and phenotypic differentiation of Arabidopsis thaliana along altitudinal gradients in the North Italian Alps

Altitudinal gradients in mountain regions are short‐range clines of different environmental parameters such as temperature or radiation. We investigated genomic and phenotypic signatures of adaptation to such gradients in five Arabidopsis thaliana populations from the North Italian Alps that originated from 580 to 2350 m altitude by resequencing pools of 19–29 individuals from each population. The sample includes two pairs of low‐ and high‐altitude populations from two different valleys. High‐altitude populations showed a lower nucleotide diversity and negative Tajima's D values and were more closely related to each other than to low‐altitude populations from the same valley. Despite their close geographic proximity, demographic analysis revealed that low‐ and high‐altitude populations split between 260 000 and 15 000 years before present. Single nucleotide polymorphisms whose allele frequencies were highly differentiated between low‐ and high‐altitude populations identified genomic regions of up to 50 kb length where patterns of genetic diversity are consistent with signatures of local selective sweeps. These regions harbour multiple genes involved in stress response. Variation among populations in two putative adaptive phenotypic traits, frost tolerance and response to light/UV stress was not correlated with altitude. Taken together, the spatial distribution of genetic diversity reflects a potentially adaptive differentiation between low‐ and high‐altitude populations, whereas the phenotypic differentiation in the two traits investigated does not. It may resemble an interaction between adaptation to the local microhabitat and demographic history influenced by historical glaciation cycles, recent seed dispersal and genetic drift in local populations.     

Identification of genomic regions associated with phenotypic variation between dog breeds using selection mapping

The extraordinary phenotypic diversity of dog breeds has been sculpted by a unique population history accompanied by selection for novel and desirable traits. Here we perform a comprehensive analysis using multiple test statistics to identify regions under selection in 509 dogs from 46 diverse breeds using a newly developed high-density genotyping array consisting of >170,000 evenly spaced SNPs. We first identify 44 genomic regions exhibiting extreme differentiation across multiple breeds. Genetic variation in these regions correlates with variation in several phenotypic traits that vary between breeds, and we identify novel associations with both morphological and behavioral traits. We next scan the genome for signatures of selective sweeps in single breeds, characterized by long regions of reduced heterozygosity and fixation of extended haplotypes. These scans identify hundreds of regions, including 22 blocks of homozygosity longer than one megabase in certain breeds. Candidate selection loci are strongly enriched for developmental genes. We chose one highly differentiated region, associated with body size and ear morphology, and characterized it using high-throughput sequencing to provide a list of variants that may directly affect these traits. This study provides a catalogue of genomic regions showing extreme reduction in genetic variation or population differentiation in dogs, including many linked to phenotypic variation. The many blocks of reduced haplotype diversity observed across the genome in dog breeds are the result of both selection and genetic drift, but extended blocks of homozygosity on a megabase scale appear to be best explained by selection. Further elucidation of the variants under selection will help to uncover the genetic basis of complex traits and disease.     

Matriline effects on metamorphic traits in a natural system in the European common frog (Rana temporaria)

Successful reproduction is an important determinant of the fitness of an individual and of the dynamics of populations. Offspring of the European common frog (Rana temporaria) exhibit a high degree of variability in metamorphic traits. However, environmental factors alone cannot explain this phenotypic variability, and the influence of genetic factors remains to be determined. Here, we tested whether the maternal genotype influences developmental time, body size, and body condition of offspring in a forest pond in Germany. We collected fertilized eggs from all 57 clutches deposited in the pond. We used multilocus genotypes based on seven microsatellite loci to assign metamorphosed offspring to mothers and to determine the number of fathers for a single matriline. We tested the influence of genetic effects in the same environment by comparing variability of metamorphic traits within and between full‐sib offspring grouped to matrilines and tested whether multiple paternity increases the variability of metamorphic traits in a single matriline. The variability in size and body condition was higher within matrilines than between them, which indicates that these traits are more strongly influenced by environmental effects, which are counteracting underlying genetic effects. The developmental time varied considerably between matrilines and variability increased with the effective number of fathers, suggesting an additive genetic effect of multiple paternity. Our results show that metamorphic traits are shaped by environmental as well as genetic effects.     

From QTL to QTN: Candidate Gene Set Approach and a Case Study in Porcine IGF1-FoxO Pathway

Unraveling the genetic background of economic traits is a major goal in modern animal genetics and breeding. Both candidate gene analysis and QTL mapping have previously been used for identifying genes and chromosome regions related to studied traits. However, most of these studies may be limited in their ability to fully consider how multiple genetic factors may influence a particular phenotype of interest. If possible, taking advantage of the combined effect of multiple genetic factors is expected to be more powerful than analyzing single sites, as the joint action of multiple loci within a gene or across multiple genes acting in the same gene set will likely have a greater influence on phenotypic variation. Thus, we proposed a pipeline of gene set analysis that utilized information from multiple loci to improve statistical power. We assessed the performance of this approach by both simulated and a real IGF1-FoxO pathway data set. The results showed that our new method can identify the association between genetic variation and phenotypic variation with higher statistical power and unravel the mechanisms of complex traits in a point of gene set. Additionally, the proposed pipeline is flexible to be extended to model complex genetic structures that include the interactions between different gene sets and between gene sets and environments.     

Direct and indirect induction of a compensatory phenotype that alleviates the costs of an inducible defense

Organisms often exhibit phenotypic plasticity in multiple traits in response to impending environmental change. Multiple traits phenotypic plasticity is complex syndrome brought on by causal relations in ecological and physiological context. Larvae of the salamander Hynobius retardatus exhibit inducible phenotypic plasticity of two traits, when at risk of predation by dragonfly larvae. One induced phenotype is an adaptive defense behaviour, i.e., stasis at the bottom of water column, directly triggered by the predation risk. Another one is a compensatory phenotype, i.e., enlarged external gills, for an unavoidable cost (hypoxia) associated with the induced defense. We identified two ways by which this compensatory phenotype could be induced. The compensatory phenotype is induced in response to not only the associated hypoxic conditions resulting from the induced defense but also the most primary but indirect cause, presence of the predator.