Drosophila Genetic Reference Panel Lines Research Articles

Drosophila Toxicogenomics: genetic variation and sexual dimorphism in susceptibility to 4-Methylimidazole

Background4-methylimidazole is a ubiquitous and potentially carcinogenic environmental toxicant. Genetic factors that contribute to variation in susceptibility to its toxic effects are challenging to assess in human populations. We used the Drosophila melanogaster Genetic Reference Panel (DGRP), a living library of natural genetic variation, to identify genes with human orthologs associated with variation in susceptibility to 4-methylimidazole.ResultsWe screened 204 DGRP lines for survival following 24-hour exposure to 4-methylimidazole. We found extensive genetic variation for survival, with a broad sense heritability of 0.82; as well as genetic variation in sexual dimorphism, with a cross-sex genetic correlation of 0.59. Genome-wide association analyses identified a total of 241 candidate molecular polymorphisms in or near 273 unique genes associated with survival. These polymorphisms had either sex-specific or sex-antagonistic effects, and most had putative regulatory effects. We generated interaction networks using these candidate genes as inputs and computationally recruited genes with known physical or genetic interactions. The network genes were significantly over-represented for gene ontology terms involving all aspects of development (including nervous system development) and cellular and organismal functions as well as canonical signaling pathways, and most had human orthologs.ConclusionsThe genetic basis of variation in sensitivity to acute exposure to 4-methylimidazole in Drosophila is attributable to variation in genes and networks of genes known for their effects on multiple developmental and cellular processes, including possible neurotoxicity. Given evolutionary conservation of the underlying genes and pathways, these insights may be applicable to humans.

A genome-wide association study implicates the olfactory system in Drosophila melanogaster diapause-associated lifespan extension and fecundity.

The effects of environmental stress on animal life are gaining importance with climate change. Diapause is a dormancy program that occurs in response to an adverse environment, followed by resumption of development and reproduction upon the return of favorable conditions. Diapause is a complex trait, so we leveraged the Drosophila genetic reference panel (DGRP) lines and conducted a Genome-Wide Association Study (GWAS) to characterize the genetic basis of diapause. We assessed post-diapause and non-diapause fecundity across 193 DGRP lines. GWAS revealed 546 genetic variants, encompassing single nucleotide polymorphisms, insertions and deletions associated with post-diapause fecundity. We identified 291 candidate diapause-associated genes, 40 of which had previously been associated with diapause. 89 of the candidates were associated with more than one SNP. Gene network analysis indicated that the diapause-associated genes were primarily linked to neuronal and reproductive system development. Similarly, comparison with results from other fly GWAS revealed the greatest overlap with olfactory-behavior-associated and fecundity-and-lifespan-associated genes. An RNAi screen of the top candidates identified two neuronal genes, Dip-γ and Scribbler, to be required during recovery for post-diapause fecundity. We complemented the genetic analysis with a test of which neurons are required for successful diapause. We found that although amputation of the antenna had little to no effect on non-diapause lifespan, it reduced diapause lifespan and postdiapause fecundity. We further show that olfactory receptor neurons and temperature-sensing neurons are required for successful recovery from diapause. Our results provide insights into the molecular, cellular, and genetic basis of adult reproductive diapause in Drosophila .

Diurnal variation in genetic parameters for locomotor activity in Drosophila melanogaster assessed under natural thermal conditions.

In nature, organisms are exposed to variable and occasionally stressful environmental conditions. Responses to diurnal and seasonal fluctuations, such as temperature and food accessibility, involve adaptive behavioural and physiological changes. While much work has been done on understanding the genetic architecture and evolutionary potential of stress tolerance traits under constant thermal conditions, there has been less focus on the quantitative genetic background in variable environments. In this study, we use the Drosophila Genetic Reference Panel (DGRP) to investigate the locomotor activity, a key behavioural trait, under variable natural thermal conditions during the summer in a temperate environment. Male flies from 100 DGRP lines were exposed to natural thermal and light conditions in Drosophila activity monitors across three experimental days. We found that activity was highly temperature and time dependent and varied between lines both within and between days. Furthermore, we observed variation in genetic and environmental variance components, with low to moderate estimates of the heritability for locomotor activity, consistently peaking in the afternoons. Moreover, we showed that the estimated genetic correlations of locomotor activity between two time points decreased, as the absolute differences in ambient temperature increased. In conclusion, we find that the genetic background for locomotor activity is environment specific, and we conclude that more variable and unpredictable future temperatures will likely have a strong impact on the evolutionary trajectories of behavioural traits in ectotherms.

Lissencephaly-1 mutations enhance traumatic brain injury outcomes in Drosophila.

Traumatic brain injury (TBI) outcomes vary greatly among individuals, but most of the variation remains unexplained. Using a Drosophila melanogaster TBI model and 178 genetically diverse lines from the Drosophila Genetic Reference Panel (DGRP), we investigated the role that genetic variation plays in determining TBI outcomes. Following injury at 20-27 days old, DGRP lines varied considerably in mortality within 24 h ("early mortality"). Additionally, the disparity in early mortality resulting from injury at 20-27 vs 0-7 days old differed among DGRP lines. These data support a polygenic basis for differences in TBI outcomes, where some gene variants elicit their effects by acting on aging-related processes. Our genome-wide association study of DGRP lines identified associations between single nucleotide polymorphisms in Lissencephaly-1 (Lis-1) and Patronin and early mortality following injury at 20-27 days old. Lis-1 regulates dynein, a microtubule motor required for retrograde transport of many cargoes, and Patronin protects microtubule minus ends against depolymerization. While Patronin mutants did not affect early mortality, Lis-1 compound heterozygotes (Lis-1x/Lis-1y) had increased early mortality following injury at 20-27 or 0-7 days old compared with Lis-1 heterozygotes (Lis-1x/+), and flies that survived 24 h after injury had increased neurodegeneration but an unaltered lifespan, indicating that Lis-1 affects TBI outcomes independently of effects on aging. These data suggest that Lis-1 activity is required in the brain to ameliorate TBI outcomes through effects on axonal transport, microtubule stability, and other microtubule proteins, such as tau, implicated in chronic traumatic encephalopathy, a TBI-associated neurodegenerative disease in humans.

Regulation at Drosophila's Malic Enzyme highlights the complexity of transvection and its sensitivity to genetic background.

Transvection, a type of trans-regulation of gene expression in which regulatory elements on one chromosome influence elements on a paired homologous chromosome, is itself a complex biological phenotype subject to modification by genetic background effects. However, relatively few studies have explored how transvection is affected by distal genetic variation, perhaps because it is strongly influenced by local regulatory elements and chromosomal architecture. With the emergence of the "hub" model of transvection and a series of studies showing variation in transvection effects, it is becoming clear that genetic background plays an important role in how transvection influences gene transcription. We explored the effects of genetic background on transvection by performing two independent genome wide association studies (GWASs) using the Drosophila genetic reference panel (DGRP) and a suite of Malic enzyme (Men) excision alleles. We found substantial variation in the amount of transvection in the 149 DGRP lines used, with broad-sense heritability of 0.89 and 0.84, depending on the excision allele used. The specific genetic variation identified was dependent on the excision allele used, highlighting the complex genetic interactions influencing transvection. We focussed primarily on genes identified as significant using a relaxed P-value cutoff in both GWASs. The most strongly associated genetic variant mapped to an intergenic single nucleotide polymorphism (SNP), located upstream of Tiggrin (Tig), a gene that codes for an extracellular matrix protein. Variants in other genes, such transcription factors (CG7368 and Sima), RNA binding proteins (CG10418, Rbp6, and Rig), enzymes (AdamTS-A, CG9743, and Pgant8), proteins influencing cell cycle progression (Dally and Eip63E) and signaling proteins (Atg-1, Axo, Egfr, and Path) also associated with transvection in Men. Although not intuitively obvious how many of these genes may influence transvection, some have been previously identified as promoting or antagonizing somatic homolog pairing. These results identify several candidate genes to further explore in the understanding of transvection in Men and in other genes regulated by transvection. Overall, these findings highlight the complexity of the interactions involved in gene regulation, even in phenotypes, such as transvection, that were traditionally considered to be primarily influenced by local genetic variation.

Modulation of the Drosophila transcriptome by developmental exposure to alcohol.

BackgroundPrenatal exposure to ethanol can cause fetal alcohol spectrum disorder (FASD), a prevalent, preventable pediatric disorder. Identifying genetic risk alleles for FASD is challenging since time, dose, and frequency of exposure are often unknown, and manifestations of FASD are diverse and evident long after exposure. Drosophila melanogaster is an excellent model to study the genetic basis of the effects of developmental alcohol exposure since many individuals of the same genotype can be reared under controlled environmental conditions.ResultsWe used 96 sequenced, wild-derived inbred lines from the Drosophila melanogaster Genetic Reference Panel (DGRP) to profile genome-wide transcript abundances in young adult flies that developed on ethanol-supplemented medium or standard culture medium. We found substantial genetic variation in gene expression in response to ethanol with extensive sexual dimorphism. We constructed sex-specific genetic networks associated with alcohol-dependent modulation of gene expression that include protein-coding genes, Novel Transcribed Regions (NTRs, postulated to encode long non-coding RNAs) and female-specific coordinated regulation of snoRNAs that regulate pseudouridylation of ribosomal RNA. We reared DGRP lines which showed extreme upregulation or downregulation of snoRNA expression during developmental alcohol exposure on standard or ethanol supplemented medium and demonstrated that developmental exposure to ethanol has genotype-specific effects on adult locomotor activity and sleep.ConclusionsThere is significant and sex-specific natural genetic variation in the transcriptional response to developmental exposure to ethanol in Drosophila that comprises networks of genes affecting nervous system development and ethanol metabolism as well as networks of regulatory non-coding RNAs.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12864-022-08559-9.

Generation of isofemale lines from hypoxia tolerant Drosophila melanogaster population

Hypoxia is a common pathological element in many human diseases. Understanding the mechanisms that underlie hypoxia tolerance or susceptibility is essential for developing novel strategies for treatment or prevention. Through experimental evolution, we obtained a Drosophila melanogaster population that can live perpetually in severe, normally lethal, hypoxic environments. We generated a Panel of Low Oxygen Tolerant (PLOT) isofemale lines (total 79 lines) from this hypoxia tolerant population to study or identify the specific genetic mechanisms that are responsible for the hypoxia tolerant trait. These isofemale lines showed significantly higher hypoxia tolerance as compared to their wildtype parental isofemale lines and the wildtype DGRP (Drosophila melanogaster Genetic Reference Panel) isofemale lines. For example, 95% of the PLOT lines showed hypoxia tolerance with eclosion rate >50% at 5% O2 level. In contrast, none of the parental lines and only 15% of the DGRP lines had similar eclosion rates under the same hypoxic condition. Furthermore, under the lethal condition with 4% O2, our PLOT lines exhibited a wide range of eclosion rate from 1% to 80% with a clear pattern of Gaussian‐Distribution, demonstrating that these PLOT flies may provide us a unique opportunity to study the molecular basis of stress‐directed evolution, such as the role of genetic variations in the protein coding regions and functional DNA elements in hypoxia adaptation. These PLOT lines can also be used to study phenotypic plasticity as well as genetic‐epigenetic interactions in hypoxic, or other stressful, environments.

The metabolome as a link in the genotype-phenotype map for peroxide resistance in the fruit fly, Drosophila melanogaster

BackgroundGenetic association studies that seek to explain the inheritance of complex traits typically fail to explain a majority of the heritability of the trait under study. Thus, we are left with a gap in the map from genotype to phenotype. Several approaches have been used to fill this gap, including those that attempt to map endophenotype such as the transcriptome, proteome or metabolome, that underlie complex traits. Here we used metabolomics to explore the nature of genetic variation for hydrogen peroxide (H2O2) resistance in the sequenced inbred Drosophila Genetic Reference Panel (DGRP).ResultsWe first studied genetic variation for H2O2 resistance in 179 DGRP lines and along with identifying the insulin signaling modulator u-shaped and several regulators of feeding behavior, we estimate that a substantial amount of phenotypic variation can be explained by a polygenic model of genetic variation. We then profiled a portion of the aqueous metabolome in subsets of eight ‘high resistance’ lines and eight ‘low resistance’ lines. We used these lines to represent collections of genotypes that were either resistant or sensitive to the stressor, effectively modeling a discrete trait. Across the range of genotypes in both populations, flies exhibited surprising consistency in their metabolomic signature of resistance. Importantly, the resistance phenotype of these flies was more easily distinguished by their metabolome profiles than by their genotypes. Furthermore, we found a metabolic response to H2O2 in sensitive, but not in resistant genotypes. Metabolomic data further implicated at least two pathways, glycogen and folate metabolism, as determinants of sensitivity to H2O2. We also discovered a confounding effect of feeding behavior on assays involving supplemented food.ConclusionsThis work suggests that the metabolome can be a point of convergence for genetic variation influencing complex traits, and can efficiently elucidate mechanisms underlying trait variation.

Genetic Variation and Potential for Resistance Development to the tTA Overexpression Lethal System in Insects.

Release of insect pests carrying the dominant lethal tetracycline transactivator (tTA) overexpression system has been proposed as a means for population suppression. High levels of the tTA transcription factor are thought to be toxic due to either transcriptional squelching or interference with protein ubiquitination. Here we utilized the Drosophila melanogaster Genetic Reference Panel (DGRP) to examine the influence of genetic variation on the efficacy of a female-specific tTA overexpression system. The level of female lethality between DGRP lines varied from 11 to 97% with a broad sense heritability of 0.89. A genome-wide association analysis identified 192 allelic variants associated with high or low lethality (P < 10−5), although none were significant when corrected for multiple testing. 151 of the variants fell within 108 genes that were associated with several biological processes including transcription and protein ubiquitination. In four lines with high female lethality, tTA RNA levels were similar or higher than in the parental tTA overexpression strain. In two lines with low lethality, tTA levels were about two fold lower than in the parental strain. However, in two other lines with low lethality, tTA levels were similar or approximately 30% lower. RNAseq analysis identified genes that were up or downregulated in the four low female lethal lines compared to the four high lethal lines. For example, genes associated with RNA processing and rRNA maturation were significantly upregulated in low lethal lines. Our data suggest that standing genetic variation in an insect population could provide multiple mechanisms for resistance to the tTA overexpression system.

Context-dependent genetic architecture of Drosophila life span.

Understanding the genetic basis of variation in life span is a major challenge that is difficult to address in human populations. Evolutionary theory predicts that alleles affecting natural variation in life span will have properties that enable them to persist in populations at intermediate frequencies, such as late-life–specific deleterious effects, antagonistic pleiotropic effects on early and late-age fitness components, and/or sex- and environment-specific or antagonistic effects. Here, we quantified variation in life span in males and females reared in 3 thermal environments for the sequenced, inbred lines of the Drosophila melanogaster Genetic Reference Panel (DGRP) and an advanced intercross outbred population derived from a subset of DGRP lines. Quantitative genetic analyses of life span and the micro-environmental variance of life span in the DGRP revealed significant genetic variance for both traits within each sex and environment, as well as significant genotype-by-sex interaction (GSI) and genotype-by-environment interaction (GEI). Genome-wide association (GWA) mapping in both populations implicates over 2,000 candidate genes with sex- and environment-specific or antagonistic pleiotropic allelic effects. Over 1,000 of these genes are associated with variation in life span in other D. melanogaster populations. We functionally assessed the effects of 15 candidate genes using RNA interference (RNAi): all affected life span and/or micro-environmental variance of life span in at least one sex and environment and exhibited sex-and environment-specific effects. Our results implicate novel candidate genes affecting life span and suggest that variation for life span may be maintained by variable allelic effects in heterogeneous environments.

Cis-regulatory variation modulates susceptibility to enteric infection in the Drosophila genetic reference panel

BackgroundResistance to enteric pathogens is a complex trait at the crossroads of multiple biological processes. We have previously shown in the Drosophila Genetic Reference Panel (DGRP) that resistance to infection is highly heritable, but our understanding of how the effects of genetic variants affect different molecular mechanisms to determine gut immunocompetence is still limited.ResultsTo address this, we perform a systems genetics analysis of the gut transcriptomes from 38 DGRP lines that were orally infected with Pseudomonas entomophila. We identify a large number of condition-specific, expression quantitative trait loci (local-eQTLs) with infection-specific ones located in regions enriched for FOX transcription factor motifs. By assessing the allelic imbalance in the transcriptomes of 19 F1 hybrid lines from a large round robin design, we independently attribute a robust cis-regulatory effect to only 10% of these detected local-eQTLs. However, additional analyses indicate that many local-eQTLs may act in trans instead. Comparison of the transcriptomes of DGRP lines that were either susceptible or resistant to Pseudomonas entomophila infection reveals nutcracker as the only differentially expressed gene. Interestingly, we find that nutcracker is linked to infection-specific eQTLs that correlate with its expression level and to enteric infection susceptibility. Further regulatory analysis reveals one particular eQTL that significantly decreases the binding affinity for the repressor Broad, driving differential allele-specific nutcracker expression.ConclusionsOur collective findings point to a large number of infection-specific cis- and trans-acting eQTLs in the DGRP, including one common non-coding variant that lowers enteric infection susceptibility.

Mitochondrial haplotypes affect metabolic phenotypes in the Drosophila Genetic Reference Panel.

The nature and extent of mitochondrial DNA variation in a population and how it affects traits is poorly understood. Here we resequence the mitochondrial genomes of 169 Drosophila Genetic Reference Panel lines, identifying 231 variants that stratify along 12 mitochondrial haplotypes. We identify 1,845 cases of mitonuclear allelic imbalances, thus implying that mitochondrial haplotypes are reflected in the nuclear genome. However, no major fitness effects are associated with mitonuclear imbalance, suggesting that such imbalances reflect population structure at the mitochondrial level rather than genomic incompatibilities. Although mitochondrial haplotypes have no direct impact on mitochondrial respiration, some haplotypes are associated with stress- and metabolism-related phenotypes, including food intake in males. Finally, through reciprocal swapping of mitochondrial genomes, we demonstrate that a mitochondrial haplotype associated with high food intake can rescue a low food intake phenotype. Together, our findings provide new insight into population structure at the mitochondrial level and point to the importance of incorporating mitochondrial haplotypes in genotype-phenotype relationship studies.

Intraspecific Genetic Variation for Lead-Induced Changes in Reproductive Strategies.

We aimed to identify genetic variation in the response of reproductive behaviors to lead (Pb2+) exposure. We reared a subset of the Drosophila Genetic Reference Panel (DGRP) inbred lines on control or Pb-treated (500μM PbAc) medium and tested for differences in copulation latency, copulation duration, and fecundity. Pb exposure decreased fecundity (p < 0.05) and increased copulation duration (p < 0.05) across DGRP lines. We found intraspecific genetic variation in latency, duration, and fecundity in both control and Pb-treated flies, with heritability ranging from 0.45 to 0.80. We found a significant genotype-by-environment interaction for copulation duration (p < 0.05). Genetic correlation matrices revealed significant genetic variation in common between control and Pb-treated flies for each trait (p < 0.05). Our results indicate that intraspecific genetic variation plays a role in Pb susceptibility and emphasize the importance of considering the impacts of variation in susceptibility to Pb pollution.

The Drosophila Gene Sulfateless Modulates Autism-Like Behaviors.

Major challenges to identifying genes that contribute to autism spectrum disorder (ASD) risk include the availability of large ASD cohorts, the contribution of many genes overall, and small effect sizes attributable to common gene variants. An alternative approach is to use a model organism to detect alleles that impact ASD-relevant behaviors and ask whether homologous human genes infer ASD risk. Here we utilized the Drosophila genetic reference panel (DGRP) as a tool to probe for perturbation in naturally occurring behaviors in Drosophila melanogaster that are analogous to three behavior domains: impaired social communication, social reciprocity and repetitive behaviors or restricted interests. Using 40 of the available DGRP lines, we identified single nucleotide polymorphisms (SNPs) in or near genes controlling these behavior domains, including ASD gene orthologs (neurexin 4 and neuroligin 2), an intellectual disability (ID) gene homolog (kirre), and a gene encoding a heparan sulfate (HS) modifying enzyme called sulfateless (sfl). SNPs in sfl were associated with all three ASD-like behaviors. Using RNAi knock-down of neuronal sfl expression, we observed significant changes in expressive and receptive communication during mating, decreased grooming behavior, and increased social spacing. These results suggest a role for HS proteoglycan synthesis and/or modification in normal social communication, repetitive behavior, and social interaction in flies. Finally, using the DGRP to directly identify genetic effects relevant to a neuropsychiatric disorder further demonstrates the utility of the Drosophila system in the discovery of genes relevant to human disease.

Lisinopril Preserves Physical Resilience and Extends Life Span in a Genotype-Specific Manner in Drosophila melanogaster.

Physical resiliency declines with age and comorbid conditions. In humans, angiotensin-converting enzyme (ACE) has been associated with attenuation of the decline in physical performance with age. ACE-inhibitor compounds, commonly prescribed for hypertension, often have beneficial effects on physical performance however the generality of these effects are unclear. Here, we tested the effects of the ACE-inhibitor Lisinopril on life span, and age-specific speed, endurance, and strength using three genotypes of the Drosophila melanogaster Genetic Reference Panel. We show that age-related decline in physical performance and survivorship varies with genetic background. Lisinopril treatment increased mean life span in all Drosophila Genetic Reference Panel lines, but its effects on life span, speed, endurance, and strength depended on genotype. We show that genotypes with increased physical performance on Lisinopril treatment experienced reduced age-related protein aggregation in muscle. Knockdown of skeletal muscle-specific Ance, the Drosophila ortholog of ACE, abolished the effects of Lisinopril on life span, implying a role for skeletal muscle Ance in survivorship. Using transcriptome profiling, we identified genes involved in stress response that showed expression changes associated with genotype and age-dependent responsiveness to Lisinopril. Our results demonstrate that Ance is involved in physical decline and demonstrate genetic variation in phenotypic responses to an ACE inhibitor.

Genome-Wide Association Study of Circadian Behavior in Drosophila melanogaster

Circadian rhythms influence physiological processes from sleep–wake cycles to body temperature and are controlled by highly conserved cycling molecules. Although the mechanistic basis of the circadian clock has been known for decades, the extent to which circadian rhythms vary in nature and the underlying genetic basis for that variation is not well understood. We measured circadian period (Ʈ) and rhythmicity index in the Drosophila Genetic Reference Panel (DGRP) and observed extensive genetic variation in both. Seven DGRP lines had sexually dimorphic arrhythmicity and one line had an exceptionally long Ʈ. Genome-wide analyses identified 584 polymorphisms in 268 genes. We observed differences among transcripts for nine genes predicted to interact among themselves and canonical clock genes in the long period line and a control. Mutations/RNAi knockdown targeting these genes also affected circadian behavior. Our observations reveal that complex genetic interactions influence high levels of variation in circadian phenotypes.

A Cyclin E Centered Genetic Network Contributes to Alcohol-Induced Variation in Drosophila Development.

Prenatal exposure to ethanol causes a wide range of adverse physiological, behavioral and cognitive consequences. However, identifying allelic variants and genetic networks associated with variation in susceptibility to prenatal alcohol exposure is challenging in human populations, since time and frequency of exposure and effective dose cannot be determined quantitatively and phenotypic manifestations are diverse. Here, we harnessed the power of natural variation in the Drosophila melanogaster Genetic Reference Panel (DGRP) to identify genes and genetic networks associated with variation in sensitivity to developmental alcohol exposure. We measured development time from egg to adult and viability of 201 DGRP lines reared on regular or ethanol- supplemented medium and identified polymorphisms associated with variation in susceptibility to developmental ethanol exposure. We also documented genotype-dependent variation in sensorimotor behavior after developmental exposure to ethanol using the startle response assay in a subset of 39 DGRP lines. Genes associated with development, including development of the nervous system, featured prominently among genes that harbored variants associated with differential sensitivity to developmental ethanol exposure. Many of them have human orthologs and mutational analyses and RNAi targeting functionally validated a high percentage of candidate genes. Analysis of genetic interaction networks identified Cyclin E (CycE) as a central, highly interconnected hub gene. Cyclin E encodes a protein kinase associated with cell cycle regulation and is prominently expressed in ovaries. Thus, exposure to ethanol during development of Drosophila melanogaster might serve as a genetic model for translational studies on fetal alcohol spectrum disorder.

Charting the genotype-phenotype map: lessons from the Drosophila melanogaster Genetic Reference Panel.

Understanding the genetic architecture (causal molecular variants, their effects, and frequencies) of quantitative traits is important for precision agriculture and medicine and predicting adaptive evolution, but is challenging in most species. The Drosophila melanogaster Genetic Reference Panel (DGRP) is a collection of 205 inbred strains with whole genome sequences derived from a single wild population in Raleigh, NC, USA. The large amount of quantitative genetic variation, lack of population structure, and rapid local decay of linkage disequilibrium in the DGRP and outbred populations derived from DGRP lines present a favorable scenario for performing genome-wide association (GWA) mapping analyses to identify candidate causal genes, polymorphisms, and pathways affecting quantitative traits. The many GWA studies utilizing the DGRP have revealed substantial natural genetic variation for all reported traits, little evidence for variants with large effects but enrichment for variants with low P-values, and a tendency for lower frequency variants to have larger effects than more common variants. The variants detected in the GWA analyses rarely overlap those discovered using mutagenesis, and often are the first functional annotations of computationally predicted genes. Variants implicated in GWA analyses typically have sex-specific and genetic background-specific (epistatic) effects, as well as pleiotropic effects on other quantitative traits. Studies in the DGRP reveal substantial genetic control of environmental variation. Taking account of genetic architecture can greatly improve genomic prediction in the DGRP. These features of the genetic architecture of quantitative traits are likely to apply to other species, including humans. WIREs Dev Biol 2018, 7:e289. doi: 10.1002/wdev.289 This article is categorized under: Invertebrate Organogenesis > Flies.

0011 ARTIFICIAL SELECTION FOR LONG AND SHORT SLEEP DURATION IN DROSOPHILA MAPS TO BROAD DEVELOPMENTAL AND SIGNALING PATHWAY GENES

Although many candidate genes have been identified for sleep, the extent to which naturally occurring polymorphisms can alter sleep duration is unknown. Here we applied an artificial selection scheme to a natural population of flies in order to determine how far night sleep duration could be driven up or down. We created an outbred population from ten Drosophila Genetic Reference Panel lines with variable sleep duration. From this outbred population, we created two short and two long-sleeping populations. We measured sleep in 100 males and 100 females of each population every generation, choosing the most extreme 25 sleepers of each sex as parents for the next generation. We maintained two additional populations as unselected controls. We used whole-genome sequences across seven generations to identify allelic variants responsible for the phenotypic changes. After 13 generations of selection, long sleepers averaged 9.97 hours more nightly sleep than short sleepers. Mean 24-hour sleep was 198.9 ± 9.5 minutes in short sleepers, comparable to severe short-sleeping flies engineered with single-gene mutations; yet these flies were created from combinations of naturally occurring alleles. Selection for night sleep altered other sleep traits such as day sleep duration and night average bout length, but the numbers of night bouts were reduced in both long and short sleepers, increasing the consolidation of sleep. Flies from these populations also had normal lifespan, suggesting that there is little physiological consequence to being a natural long or short sleeper. Whole-genome sequences revealed thousands of changes in polymorphic allele frequencies between any two generations of selection. However, regressing allele frequency change across generations reduced the number of candidate polymorphisms to 126, implicating 80 candidate genes. Many of these genes could be connected via known physical and genetic interactions with several conserved developmental and signaling pathways such as the EGFR and Wnt pathways. Sleep duration can be driven to very high or low levels, suggesting that extreme short or long sleepers may exist in nature. Highly conserved broad developmental and signaling pathways influence natural variation in sleep duration. Intramural Research Program, National Institutes of Health, NHLBI.

α-amanitin resistance in Drosophila melanogaster: A genome-wide association approach.

We investigated the mechanisms of mushroom toxin resistance in the Drosophila Genetic Reference Panel (DGRP) fly lines, using genome-wide association studies (GWAS). While Drosophila melanogaster avoids mushrooms in nature, some lines are surprisingly resistant to α-amanitin—a toxin found solely in mushrooms. This resistance may represent a pre-adaptation, which might enable this species to invade the mushroom niche in the future. Although our previous microarray study had strongly suggested that pesticide-metabolizing detoxification genes confer α-amanitin resistance in a Taiwanese D. melanogaster line Ama-KTT, none of the traditional detoxification genes were among the top candidate genes resulting from the GWAS in the current study. Instead, we identified Megalin, Tequila, and widerborst as candidate genes underlying the α-amanitin resistance phenotype in the North American DGRP lines, all three of which are connected to the Target of Rapamycin (TOR) pathway. Both widerborst and Tequila are upstream regulators of TOR, and TOR is a key regulator of autophagy and Megalin-mediated endocytosis. We suggest that endocytosis and autophagy of α-amanitin, followed by lysosomal degradation of the toxin, is one of the mechanisms that confer α-amanitin resistance in the DGRP lines.