From GWAS to signal validation: An approach for estimating genetic effects while preserving genomic context.

Validating associations between genotypic and phenotypic variation remains a challenge, despite advancements in association studies. Common approaches for signal validation rely on gene-level perturbations, such as loss-of-function mutations or RNAi, which test the effect of genetic modifications usually not observed in nature. CRISPR-based methods can validate associations at the SNP level, but have significant drawbacks, including resulting off-target effects and being both time-consuming and expensive. Both approaches usually modify the genome of a single genetic background, limiting the generalizability of experiments. To address these challenges, we present a simple, low-cost experimental scheme for validating genetic associations at the SNP level in outbred populations. The approach involves genotyping live outbred individuals at a focal SNP, crossing homozygous individuals with the same genotype at that locus, and contrasting phenotypes across resulting synthetic outbred populations. We tested this method in Drosophila melanogaster, measuring the longevity effects of a polymorphism at a naturally-segregating cis-eQTL for the midway gene. Our results demonstrate the utility of this method in SNP-level validation of naturally occurring genetic variation regulating complex traits. This method provides a bridge between the statistical discovery of genotype-phenotype associations and their validation in the natural context of heterogeneous genomic contexts.

Socioeconomic status effects on health vary between rural and urban Turkana.

BACKGROUND AND OBJECTIVES: Understanding the social determinants of health is a major goal in evolutionary biology and human health research. Low socioeconomic status (often operationalized as absolute material wealth) is consistently associated with chronic stress, poor health and premature death in high-income countries. However, the degree to which wealth gradients in health are universal-or are instead made even steeper under contemporary, post-industrial conditions-remains poorly understood. METHODOLOGY: We quantified absolute material wealth and several health outcomes among a population of traditional pastoralists, the Turkana of northwest Kenya, who are currently transitioning toward a more urban, market-integrated lifestyle. We assessed whether wealth associations with health differed in subsistence-level versus urban contexts. We also explored the causes and consequences of wealth-health associations by measuring serum cortisol, potential sociobehavioral mediators in early life and adulthood, and adult reproductive success (number of surviving offspring). RESULTS: Higher socioeconomic status and greater material wealth predicts better self-reported health and more offspring in traditional pastoralist Turkana, but worse cardiometabolic health and fewer offspring in urban Turkana. We do not find robust evidence for either direct biological mediators (cortisol) or indirect sociobehavioral mediators (e.g. adult diet or health behaviors, early life experiences) of wealth-health relationships in either context. CONCLUSIONS AND IMPLICATIONS: While social gradients in health are well-established in humans and animals across a variety of socioecological contexts, we show that the relationship between wealth and health can vary within a single population. Our findings emphasize that changes in economic and societal circumstances may directly alter how, why and under what conditions socioeconomic status predicts health. LAY SUMMARY: High socioeconomic status predicts better health and more offspring in traditional Turkana pastoralists, but worse health and fewer offspring in individuals of the same group living in urban areas. Together, our study shows that under different economic and societal circumstances, wealth effects on health may manifest in very different ways.

Drosophila DNA/RNA methyltransferase contributes to robust host defense in aging animals by regulating sphingolipid metabolism.

Drosophila methyltransferase (Mt2) has been implicated in the methylation of both DNA and tRNA. In this study, we demonstrate that loss of Mt2 activity leads to an age-dependent decline of immune function in the adult fly. A newly eclosed adult has mild immune defects that are exacerbated in a 15 day old Mt2-/- fly. The age-dependent effects appear to be systemic, including disturbances in lipid metabolism, changes in cell shape of hemocytes and significant fold-changes in levels of transcripts related to host defense. Lipid imbalance, as measured by quantitative lipidomics, correlates with immune dysfunction, with high levels of immunomodulatory lipids, sphingosine-1-phosphate (S1P) and ceramides, along with low levels of storage lipids. Activity assays on fly lysates confirm the age-dependent increase in S1P and concomitant reduction of S1P lyase activity. We hypothesize that Mt2 functions to regulate genetic loci such as S1P lyase and this regulation is essential for robust host defense as the animal ages. Our study uncovers novel links between age--dependent Mt2 function, innate immune response and lipid homeostasis.

Presence of DNA methyltransferase activity and CpC methylation in Drosophila melanogaster.

Drosophila melanogaster lacks DNMT1/DNMT3 based methylation machinery. Despite recent reports confirming the presence of low DNA methylation in Drosophila; little is known about the methyltransferase. Therefore, in this study, we have aimed to investigate the possible functioning of DNA methyltransferase in Drosophila. The 14 K oligo microarray slide was incubated with native cell extract from adult Drosophila to check the presence of the methyltransferase activity. After incubation under appropriate conditions, the methylated oligo sequences were identified by the binding of anti 5-methylcytosine monoclonal antibody. The antibody bound to the methylated oligos was detected using Cy3 labeled secondary antibody. Methylation sensitive restriction enzyme mediated PCR was used to assess the methylation at a few selected loci identified on the array. It could be seen that a few of the total oligos got methylated under the assay conditions. Analysis of methylated oligo sequences provides evidence for the presence of de novo methyltransferase activity and allows identification of its sequence specificity in adult Drosophila. With the help of methylation sensitive enzymes we could detect presence of CpC methylation in the selected genomic regions. This study reports presence of an active DNA methyltransferase in adult Drosophila, which exhibits sequence specificity confirmed by presence of asymmetric methylation at corresponding sites in the genomic DNA. It also provides an innovative approach to investigate methylation specificity of a native methyltransferase.