Listeria monocytogenes Infection Causes Metabolic Shifts in Drosophila melanogaster

Immunity and metabolism are intimately linked; manipulating metabolism, either through diet or genetics, has the power to alter survival during infection. However, despite metabolism''s powerful ability to alter the course of infections, little is known about what being “sick” means metabolically. Here we describe the metabolic changes occurring in a model system when Listeria monocytogenes causes a lethal infection in Drosophila melanogaster. L. monocytogenes infection alters energy metabolism; the flies gradually lose both of their energy stores, triglycerides and glycogen, and show decreases in both intermediate metabolites and enzyme message for the two main energy pathways, beta-oxidation and glycolysis. L. monocytogenes infection also causes enzymatic reduction in the anti-oxidant uric acid, and knocking out the enzyme uric oxidase has a complicated effect on immunity. Free amino acid levels also change during infection, including a drop in tyrosine levels which may be due to robust L. monocytogenes induced melanization.     

Parallel Geographic Variation in Drosophila melanogaster

Drosophila melanogaster, an ancestrally African species, has recently spread throughout the world, associated with human activity. The species has served as the focus of many studies investigating local adaptation relating to latitudinal variation in non-African populations, especially those from the United States and Australia. These studies have documented the existence of shared, genetically determined phenotypic clines for several life history and morphological traits. However, there are no studies designed to formally address the degree of shared latitudinal differentiation at the genomic level. Here we present our comparative analysis of such differentiation. Not surprisingly, we find evidence of substantial, shared selection responses on the two continents, probably resulting from selection on standing ancestral variation. The polymorphic inversion In(3R)P has an important effect on this pattern, but considerable parallelism is also observed across the genome in regions not associated with inversion polymorphism. Interestingly, parallel latitudinal differentiation is observed even for variants that are not particularly strongly differentiated, which suggests that very large numbers of polymorphisms are targets of spatially varying selection in this species.     

The NADPH Metabolic Network Regulates Human αB-crystallin Cardiomyopathy and Reductive Stress in Drosophila melanogaster

Dominant mutations in the alpha-B crystallin (CryAB) gene are responsible for a number of inherited human disorders, including cardiomyopathy, skeletal muscle myopathy, and cataracts. The cellular mechanisms of disease pathology for these disorders are not well understood. Among recent advances is that the disease state can be linked to a disturbance in the oxidation/reduction environment of the cell. In a mouse model, cardiomyopathy caused by the dominant CryAB^R120G missense mutation was suppressed by mutation of the gene that encodes glucose 6-phosphate dehydrogenase (G6PD), one of the cell''s primary sources of reducing equivalents in the form of NADPH. Here, we report the development of a Drosophila model for cellular dysfunction caused by this CryAB mutation. With this model, we confirmed the link between G6PD and mutant CryAB pathology by finding that reduction of G6PD expression suppressed the phenotype while overexpression enhanced it. Moreover, we find that expression of mutant CryAB in the Drosophila heart impaired cardiac function and increased heart tube dimensions, similar to the effects produced in mice and humans, and that reduction of G6PD ameliorated these effects. Finally, to determine whether CryAB pathology responds generally to NADPH levels we tested mutants or RNAi-mediated knockdowns of phosphogluconate dehydrogenase (PGD), isocitrate dehydrogenase (IDH), and malic enzyme (MEN), the other major enzymatic sources of NADPH, and we found that all are capable of suppressing CryAB^R120G pathology, confirming the link between NADP/H metabolism and CryAB.     

Population Genomics of Inversion Polymorphisms in Drosophila melanogaster

Chromosomal inversions have been an enduring interest of population geneticists since their discovery in Drosophila melanogaster. Numerous lines of evidence suggest powerful selective pressures govern the distributions of polymorphic inversions, and these observations have spurred the development of many explanatory models. However, due to a paucity of nucleotide data, little progress has been made towards investigating selective hypotheses or towards inferring the genealogical histories of inversions, which can inform models of inversion evolution and suggest selective mechanisms. Here, we utilize population genomic data to address persisting gaps in our knowledge of D. melanogaster''s inversions. We develop a method, termed Reference-Assisted Reassembly, to assemble unbiased, highly accurate sequences near inversion breakpoints, which we use to estimate the age and the geographic origins of polymorphic inversions. We find that inversions are young, and most are African in origin, which is consistent with the demography of the species. The data suggest that inversions interact with polymorphism not only in breakpoint regions but also chromosome-wide. Inversions remain differentiated at low levels from standard haplotypes even in regions that are distant from breakpoints. Although genetic exchange appears fairly extensive, we identify numerous regions that are qualitatively consistent with selective hypotheses. Finally, we show that In(1)Be, which we estimate to be ∼60 years old (95% CI 5.9 to 372.8 years), has likely achieved high frequency via sex-ratio segregation distortion in males. With deeper sampling, it will be possible to build on our inferences of inversion histories to rigorously test selective models—particularly those that postulate that inversions achieve a selective advantage through the maintenance of co-adapted allele complexes.     

The metabolome as a biomarker of aging in Drosophila melanogaster

Many biomarkers have been shown to be associated not only with chronological age but also with functional measures of biological age. In human populations, it is difficult to show whether variation in biological age is truly predictive of life expectancy, as such research would require longitudinal studies over many years, or even decades. We followed adult cohorts of 20 Drosophila Genetic Reference Panel (DGRP) strains chosen to represent the breadth of lifespan variation, obtain estimates of lifespan, baseline mortality, and rate of aging, and associate these parameters with age‐specific functional traits including fecundity and climbing activity and with age‐specific targeted metabolomic profiles. We show that activity levels and metabolome‐wide profiles are strongly associated with age, that numerous individual metabolites show a strong association with lifespan, and that the metabolome provides a biological clock that predicts not only sample age but also future mortality rates and lifespan. This study with 20 genotypes and 87 metabolites, while relatively small in scope, establishes strong proof of principle for the fly as a powerful experimental model to test hypotheses about biomarkers and aging and provides further evidence for the potential value of metabolomic profiles as biomarkers of aging.     

Genetic Architecture of Abdominal Pigmentation in Drosophila melanogaster

Pigmentation varies within and between species and is often adaptive. The amount of pigmentation on the abdomen of Drosophila melanogaster is a relatively simple morphological trait, which serves as a model for mapping the genetic basis of variation in complex phenotypes. Here, we assessed natural variation in female abdominal pigmentation in 175 sequenced inbred lines of the Drosophila melanogaster Genetic Reference Panel, derived from the Raleigh, NC population. We quantified the proportion of melanization on the two most posterior abdominal segments, tergites 5 and 6 (T5, T6). We found significant genetic variation in the proportion of melanization and high broad-sense heritabilities for each tergite. Genome-wide association studies identified over 150 DNA variants associated with the proportion of melanization on T5 (84), T6 (34), and the difference between T5 and T6 (35). Several of the top variants associated with variation in pigmentation are in tan, ebony, and bric-a-brac1, genes known to affect D. melanogaster abdominal pigmentation. Mutational analyses and targeted RNAi-knockdown showed that 17 out of 28 (61%) novel candidate genes implicated by the genome-wide association study affected abdominal pigmentation. Several of these genes are involved in developmental and regulatory pathways, chitin production, cuticle structure, and vesicle formation and transport. These findings show that genetic variation may affect multiple steps in pathways involved in tergite development and melanization. Variation in these novel candidates may serve as targets for adaptive evolution and sexual selection in D. melanogaster.     

Anent the Genomics of Spermatogenesis in Drosophila melanogaster

An appreciable fraction of the Drosophila melanogaster genome is dedicated to male fertility. One approach to characterizing this subset of the genome is through the study of male-sterile mutations. We studied the relation between vital and male-fertility genes in three large autosomal regions that were saturated for lethal and male-sterile mutations. The majority of male-sterile mutations affect genes that are exclusively expressed in males. These genes are required only for male fertility, and several mutant alleles of each such gene were encountered. A few male-sterile mutations were alleles of vital genes that are expressed in both males and females. About one-fifth of the genes in Drosophila melanogaster show male-specific expression in adults. Although some earlier studies found a paucity of genes on the X chromosome showing male-biased expression, we did not find any significant differences between the X chromosome and the autosomes either in the relative frequencies of mutations to male sterility or in the frequencies of genes with male-specific expression in adults. Our results suggest that as much as 25% of the Drosophila genome may be dedicated to male fertility.     

Quantitative Genetics of Food Intake in Drosophila melanogaster

Food intake is an essential animal activity, regulated by neural circuits that motivate food localization, evaluate nutritional content and acceptance or rejection responses through the gustatory system, and regulate neuroendocrine feedback loops that maintain energy homeostasis. Excess food consumption in people is associated with obesity and metabolic and cardiovascular disorders. However, little is known about the genetic basis of natural variation in food consumption. To gain insights in evolutionarily conserved genetic principles that regulate food intake, we took advantage of a model system, Drosophila melanogaster, in which food intake, environmental conditions and genetic background can be controlled precisely. We quantified variation in food intake among 182 inbred, sequenced lines of the Drosophila melanogaster Genetic Reference Panel (DGRP). We found significant genetic variation in the mean and within-line environmental variance of food consumption and observed sexual dimorphism and genetic variation in sexual dimorphism for both food intake traits (mean and variance). We performed genome wide association (GWA) analyses for mean food intake and environmental variance of food intake (using the coefficient of environmental variation, CV _E, as the metric for environmental variance) and identified molecular polymorphisms associated with both traits. Validation experiments using RNAi-knockdown confirmed 24 of 31 (77%) candidate genes affecting food intake and/or variance of food intake, and a test cross between selected DGRP lines confirmed a SNP affecting mean food intake identified in the GWA analysis. The majority of the validated candidate genes were novel with respect to feeding behavior, and many had mammalian orthologs implicated in metabolic diseases.     

Variation in Male Mate Choice in Drosophila melanogaster

Male mate choice has been reported in the fruit fly, Drosophila melanogaster, even though males of this species were previously thought to maximise their fitness by mating with all available females. To understand the evolution of male mate choice it is important to understand variation in male mating preferences. Two studies, using different stock populations and different methods, have reported contrasting patterns of variation in male mate choice in D. melanogaster. Two possible explanations are that there are evolved differences in each stock population or that the methods used to measure choice could have biased the results. We investigated these hypotheses here by repeating the methods used in one study in which variable male mate choice was found, using the stock population from the other study in which choice was not variable. The results showed a significant resource-independent male preference for less fecund, smaller females, which contrasts with previous observations of male mate choice. This indicates that different selection pressures between populations have resulted in evolved differences in the expression of male mate choice. It also reveals phenotypic plasticity in male mate choice in response to cues encountered in each choice environment. The results highlight the importance of variation in male mate choice, and of identifying mechanisms in order to understand the evolution of mate choice under varying ecological conditions.     

Genome-Wide Fine-Scale Recombination Rate Variation in Drosophila melanogaster

Estimating fine-scale recombination maps of Drosophila from population genomic data is a challenging problem, in particular because of the high background recombination rate. In this paper, a new computational method is developed to address this challenge. Through an extensive simulation study, it is demonstrated that the method allows more accurate inference, and exhibits greater robustness to the effects of natural selection and noise, compared to a well-used previous method developed for studying fine-scale recombination rate variation in the human genome. As an application, a genome-wide analysis of genetic variation data is performed for two Drosophila melanogaster populations, one from North America (Raleigh, USA) and the other from Africa (Gikongoro, Rwanda). It is shown that fine-scale recombination rate variation is widespread throughout the D. melanogaster genome, across all chromosomes and in both populations. At the fine-scale, a conservative, systematic search for evidence of recombination hotspots suggests the existence of a handful of putative hotspots each with at least a tenfold increase in intensity over the background rate. A wavelet analysis is carried out to compare the estimated recombination maps in the two populations and to quantify the extent to which recombination rates are conserved. In general, similarity is observed at very broad scales, but substantial differences are seen at fine scales. The average recombination rate of the X chromosome appears to be higher than that of the autosomes in both populations, and this pattern is much more pronounced in the African population than the North American population. The correlation between various genomic features—including recombination rates, diversity, divergence, GC content, gene content, and sequence quality—is examined using the wavelet analysis, and it is shown that the most notable difference between D. melanogaster and humans is in the correlation between recombination and diversity.     

Genome-Wide Association Analysis of Radiation Resistance in Drosophila melanogaster

Background

Ionizing radiation is genotoxic to cells. Healthy tissue toxicity in patients and radiation resistance in tumors present common clinical challenges in delivering effective radiation therapies. Radiation response is a complex, polygenic trait with unknown genetic determinants. The Drosophila Genetic Reference Panel (DGRP) provides a model to investigate the genetics of natural variation for sensitivity to radiation.

Methods and Findings

Radiation response was quantified in 154 inbred DGRP lines, among which 92 radiosensitive lines and 62 radioresistant lines were classified as controls and cases, respectively. A case-control genome-wide association screen for radioresistance was performed. There are 32 single nucleotide polymorphisms (SNPs) associated with radio resistance at a nominal p<10⁻⁵; all had modest effect sizes and were common variants with the minor allele frequency >5%. All the genes implicated by those SNP hits were novel, many without a known role in radiation resistance and some with unknown function. Variants in known DNA damage and repair genes associated with radiation response were below the significance threshold of p<10⁻⁵ and were not present among the significant hits. No SNP met the genome-wide significance threshold (p = 1.49×10⁻⁷), indicating a necessity for a larger sample size.

Conclusions

Several genes not previously associated with variation in radiation resistance were identified. These genes, especially the ones with human homologs, form the basis for exploring new pathways involved in radiation resistance in novel functional studies. An improved DGRP model with a sample size of at least 265 lines and ideally up to 793 lines is recommended for future studies of complex traits.     

Population Genomics of the Wolbachia Endosymbiont in Drosophila melanogaster

Wolbachia are maternally inherited symbiotic bacteria, commonly found in arthropods, which are able to manipulate the reproduction of their host in order to maximise their transmission. The evolutionary history of endosymbionts like Wolbachia can be revealed by integrating information on infection status in natural populations with patterns of sequence variation in Wolbachia and host mitochondrial genomes. Here we use whole-genome resequencing data from 290 lines of Drosophila melanogaster from North America, Europe, and Africa to predict Wolbachia infection status, estimate relative cytoplasmic genome copy number, and reconstruct Wolbachia and mitochondrial genome sequences. Overall, 63% of Drosophila strains were predicted to be infected with Wolbachia by our in silico analysis pipeline, which shows 99% concordance with infection status determined by diagnostic PCR. Complete Wolbachia and mitochondrial genomes show congruent phylogenies, consistent with strict vertical transmission through the maternal cytoplasm and imperfect transmission of Wolbachia. Bayesian phylogenetic analysis reveals that the most recent common ancestor of all Wolbachia and mitochondrial genomes in D. melanogaster dates to around 8,000 years ago. We find evidence for a recent global replacement of ancestral Wolbachia and mtDNA lineages, but our data suggest that the derived wMel lineage arose several thousand years ago, not in the 20th century as previously proposed. Our data also provide evidence that this global replacement event is incomplete and is likely to be one of several similar incomplete replacement events that have occurred since the out-of-Africa migration that allowed D. melanogaster to colonize worldwide habitats. This study provides a complete genomic analysis of the evolutionary mode and temporal dynamics of the D. melanogaster–Wolbachia symbiosis, as well as important resources for further analyses of the impact of Wolbachia on host biology.     

Ultrastructural Modifications in the Mitochondria of Hypoxia-Adapted Drosophila melanogaster

Chronic hypoxia (CH) occurs under certain physiological or pathological conditions, including in people who reside at high altitude or suffer chronic cardiovascular or pulmonary diseases. As mitochondria are the predominant oxygen-consuming organelles to generate ATP through oxidative phosphorylation in cells, their responses, through structural or molecular modifications, to limited oxygen supply play an important role in the overall functional adaptation to hypoxia. Here, we report the adaptive mitochondrial ultrastructural modifications and the functional impacts in a recently generated hypoxia-adapted Drosophila melanogaster strain that survives severe, otherwise lethal, hypoxic conditions. Using electron tomography, we discovered increased mitochondrial volume density and cristae abundance, yet also cristae fragmentation and a unique honeycomb-like structure in the mitochondria of hypoxia-adapted flies. The homeostatic levels of adenylate and energy charge were similar between hypoxia-adapted and naïve control flies and the hypoxia-adapted flies remained active under severe hypoxia as quantified by negative geotaxis behavior. The equilibrium ATP level was lower in hypoxia-adapted flies than those of the naïve controls tested under severe hypoxia that inhibited the motion of control flies. Our results suggest that the structural rearrangement in the mitochondria of hypoxia-adapted flies may be an important adaptive mechanism that plays a critical role in preserving adenylate homeostasis and metabolism as well as muscle function under chronic hypoxic conditions.     

Organically Grown Food Provides Health Benefits to Drosophila melanogaster

The “organic food” market is the fastest growing food sector, yet it is unclear whether organically raised food is nutritionally superior to conventionally grown food and whether consuming organic food bestows health benefits. In order to evaluate potential health benefits of organic foods, we used the well-characterized fruit fly Drosophila melanogaster as a model system. Fruit flies were raised on a diets consisting of extracts of either conventionally or organically raised produce (bananas, potatoes, raisins, soy beans). Flies were then subjected to a variety of tests designed to assess overall fly health. Flies raised on diets made from organically grown produce had greater fertility and longevity. On certain food sources, greater activity and greater stress resistance was additionally observed, suggesting that organic food bestows positive effects on fly health. Our data show that Drosophila can be used as a convenient model system to experimentally test potential health effects of dietary components. Using this system, we provide evidence that organically raised food may provide animals with tangible benefits to overall health.