Search

Expression quantitative trait locus detection has become increasingly important for understanding how noncoding variants contribute to disease susceptibility and complex traits. The major challenges in expression quantitative trait locus fine-mapping and causal variant discovery relate to the impact of linkage disequilibrium on signals due to one or multiple functional variants that lie within a credible set. We perform expression quantitative trait locus fine-mapping using the all-but-one approach, conditioning each signal on all others detected in an interval, on the Consortium for the Architecture of Gene Expression cohorts of microarray-based peripheral blood gene expression in 2,138 European-ancestry human adults. We contrast these results with traditional forward stepwise conditional analysis and a Bayesian localization method. All-but-one conditioning significantly modifies effect-size estimates for 51% of 2,351 expression quantitative trait locus peaks, but only modestly affects credible set size and location. On the other hand, both conditioning approaches result in unexpectedly low overlap with Bayesian credible sets, with just 57% peak concordance and between 50% and 70% SNP sharing, leading us to caution against the assumption that any one localization method is superior to another. We also cross reference our results with ATAC-seq data, cell-type-specific expression quantitative trait locus, and activity-by-contact-enhancers, leading to the proposal of a 5-tier approach to further reduce credible set sizes and prioritize likely causal variants for all known inflammatory bowel disease risk loci active in immune cells.

Systems biology is an approach to dissection of complex traits that explicitly recognizes the impact of genetic, physiological, and environmental interactions in the generation of phenotypic variation. We describe comprehensive transcriptional and metabolic profiling in Drosophila melanogaster across four diets, finding little overlap in modular architecture. Genotype and genotype-by-diet interactions are a major component of transcriptional variation (24 and 5.3% of the total variation, respectively) while there were no main effects of diet (

A major objective of genomics is to elucidate the mapping between genotypic and phenotypic space as a step toward understanding how small changes in gene function can lead to elaborate phenotypic changes. One approach that has been utilized is to examine overall patterns of covariation between phenotypic variables of interest, such as morphology, physiology, and behavior, and underlying aspects of gene activity, in particular transcript abundance on a genome-wide scale. Numerous studies have demonstrated that such patterns of covariation occur, although these are often between samples with large numbers of unknown genetic differences (different strains or even species) or perturbations of large effect (sexual dimorphism or strong loss-of-function mutations) that may represent physiological changes outside of the normal experiences of the organism. We used weak mutational perturbations in genes affecting wing development in Drosophila melanogaster that influence wing shape relative to a co-isogenic wild type. We profiled transcription of 1150 genes expressed during wing development in 27 heterozygous mutants, as well as their co-isogenic wild type and one additional wild-type strain. Despite finding clear evidence of expression differences between mutants and wild type, transcriptional profiles did not covary strongly with shape, suggesting that information from transcriptional profiling may not generally be predictive of final phenotype. We discuss these results in the light of possible attractor states of gene expression and how this would affect interpretation of covariation between transcriptional profiles and other phenotypes.

As a first step toward population and quantitative genetic analysis of neurotransmitter receptors in Drosophila melanogaster, we describe the parameters of nucleotide variation in three serotonin receptors and their association with pupal heart rate. Thirteen kilobases of DNA including the complete coding regions of 5-HT1A, 5-HT1B, and 5-HT2 were sequenced in 216 highly inbred lines extracted from two North American populations in California and North Carolina. Nucleotide and amino acid polymorphism is in the normal range for Drosophila genes and proteins, and linkage disequilibrium decays rapidly such that haplotype blocks are typically only a few SNPs long. However, intron 1 of 5-HT1A consists of two haplotypes that are at significantly different frequencies in the two populations. Neither this region of the gene nor any of the common amino acid polymorphisms in the three loci associate with either heart rate or heart rate variability. A cluster of SNPs in intron 2 of 5-HT1A, including a triallelic site, do show a highly significant interaction between genotype, sex, and population. While it is likely that a combination of weak, complex selection pressures and population structure has helped shape variation in the serotonin receptors of Drosophila, much larger sampling strategies than are currently adopted in evolutionary genetics will be required to disentangle these effects.

Assessment of the degree to which gene expression is additive and heritable has important implications for understanding the maintenance of variation, adaptation, phenotypic divergence, and the mapping of genotype onto phenotype. We used whole-genome transcript profiling using Agilent long-oligonucleotide microarrays representing 12,017 genes to demonstrate that gene transcription is pervasively nonadditive in Drosophila melanogaster. Comparison of adults of two isogenic lines and their reciprocal F1 hybrids revealed 5820 genes as significantly different between at least two of the four genotypes in either males or females or across both sexes. Strikingly, while 25% of all genes differ between the two parents, 33% differ between both F1's and the parents, averaged across sexes. However, only 5% of genes show overdominance, suggesting that heterosis for expression is rare.

An emerging issue in evolutionary genetics is whether it is possible to use gene expression profiling to identify genes that are associated with morphological, physiological, or behavioral divergence between species and whether these genes have undergone positive selection. Some of these questions were addressed in a recent study (Enard et al. 2002) of the difference in gene expression among human, chimp, and orangutan, which suggested an accelerated rate of divergence in gene expression in the human brain relative to liver. Reanalysis of the Affymetrix data set using analysis of variance methods to quantify the contributions of individuals and species to variation in expression of 12,600 genes indicates that as much as one-quarter of the genome shows divergent expression between primate species at the 5% level. The magnitude of fold change ranges from 1.2-fold up to 8-fold. Similar conclusions apply to reanalysis of Enard et al.'s (2002) parallel murine data set. However, biases inherent to short oligonucleotide microarray technology may account for some of the tissue and species effects. At high significance levels, more differences were observed in the liver than in the brain in each of the pairwise species comparisons, so it is not clear that expression divergence is accelerated in the human brain. Further, there is an apparent bias toward upregulation of gene expression in the brain in both primates and mice, whereas genes are equally likely to be up- or downregulated in the liver when these species diverge. A small subset of genes that are candidates for adaptive divergence may be identified on the basis of a high ratio of interspecific to intraspecific divergence.

Genetic variation affecting the expressivity of an amorphic allele of the homeotic gene Ultrabithorax, (Ubx1) was characterized after 11 generations of introgression into 29 different isofemale lines. Heterozygotes display a range of haploinsufficient phenotypes, from overlap with wild-type halteres to dramatic transformations such as a 50% increase in area and the presence of over 20 bristles on the anterior margin of each haltere. In both the wild-type and mutant genetic backgrounds, there is moderate genetic variance and low environmental variance/developmental asymmetry, as expected of a trait under stabilizing selection pressure. Surprisingly, there is little evidence that mutant halteres are more variable than wild-type ones, so it is unclear that haltere development is also canalized. The correlation between wild-type and Ubx haltere size is very low, indicating that interactions among modifiers of Ubx are complex, and in some cases sex-specific. The potential quantitative genetic contributions of homeotic genes to appendage morphology are discussed, noting that population-level effects of variation in key regulatory genes may be prevalent and complex but cannot be readily extrapolated to macroevolutionary diversification.

Genetic background effects contribute to the phenotypic consequences of mutations and are pervasive across all domains of life that have been examined, yet little is known about how they modify genetic systems. In part this is due to the lack of tractable model systems that have been explicitly developed to study the genetic and evolutionary consequences of background effects. In this study we demonstrate that phenotypic expressivity of the scallopedE3 (sdE3) mutation of Drosophila melanogaster is background dependent and is the result of at least one major modifier segregating between two standard lab wild-type strains. We provide evidence that at least one of the modifiers is linked to the vestigial region and demonstrate that the background effects modify the spatial distribution of known sd target genes in a genotype-dependent manner. In addition, microarrays were used to examine the consequences of genetic background effects on the global transcriptome. Expression differences between wild-type strains were found to be as large as or larger than the effects of mutations with substantial phenotypic effects, and expression differences between wild type and mutant varied significantly between genetic backgrounds. Significantly, we demonstrate that the epistatic interaction between sdE3 and an optomotor blind mutation is background dependent. The results are discussed within the context of developing a complex but more realistic view of the consequences of genetic background effects with respect to mutational analysis and studies of epistasis and cryptic genetic variation segregating in natural populations.

Wing development in Drosophila is a common model system for the dissection of genetic networks and their roles during development. In particular, the RTK and TGF-β regulatory networks appear to be involved with numerous aspects of wing development, including patterning, cell determination, growth, proliferation, and survival in the developing imaginal wing disc. However, little is known as to how subtle changes in the function of these genes may contribute to quantitative variation for wing shape, per se. In this study 50 insertional mutations, representing 43 loci in the RTK, Hedgehog, TGF-β pathways, and their genetically interacting factors were used to study the role of these networks on wing shape. To concurrently examine how genetic background modulates the effects of the mutation, each insertion was introgressed into two wild-type genetic backgrounds. Using geometric morphometric methods, it is shown that the majority of these mutations have profound effects on shape but not size of the wing when measured as heterozygotes. To examine the relationships between how each mutation affects wing shape hierarchical clustering was used. Unlike previous observations of environmental canalization, these mutations did not generally increase within-line variation relative to their wild-type counterparts. These results provide an entry point into the genetics of wing shape and are discussed within the framework of the dissection of complex phenotypes.

As part of an effort to dissect quantitative trait locus effects to the nucleotide level, association was assessed between 238 single-nucleotide and 20 indel polymorphisms spread over 11 kb of the Drosophila melanogaster Egfr locus and nine relative warp measures of wing shape. One SNP in a conserved potential regulatory site for a GAGA factor in the promoter of alternate first exon 2 approaches conservative experiment-wise significance (P < 0.00003) in the sample of 207 lines for association with the location of the crossveins in the central region of the wing. Several other sites indicate marginal association with one or more other aspects of shape. No strong effects of sex or population of origin were detected with measures of shape, but two different sites were strongly associated with overall wing size in interaction with these fixed factors. Whole-gene sequencing in very large samples, rather than selective genotyping, would appear to be the only strategy likely to be successful for detecting subtle associations in species with high polymorphism and little haplotype structure. However, these features severely limit the ability of linkage disequilibrium mapping in Drosophila to resolve quantitative effects to single nucleotides., A.P. was funded in part by awards from the American Scandinavian Foundation and NATO, and the project was funded by grants to G.G. from the David and Lucille Packard Foundation and the National Institutes of Health (R01 GM61600).

Drosophila melanogaster appears to be well suited as a model organism for quantitative pharmacogenetic analysis. A genome-wide deficiency screen for haploinsufficient effects on prepupal heart rate identified nine regions of the genome that significantly reduce (five deficiencies) or increase (four deficiencies) heart rate across a range of genetic backgrounds. Candidate genes include several neurotransmitter receptor loci, particularly monoamine receptors, consistent with results of prior pharmacological manipulations of heart rate, as well as genes associated with paralytic phenotypes. Significant genetic variation is also shown to exist for a suite of four autonomic behaviors that are exhibited spontaneously upon decapitation, namely, grooming, grasping, righting, and quivering. Overall activity levels are increased by application of particular concentrations of the drugs octopamine and nicotine, but due to high environmental variance both within and among replicate vials, the significance of genetic variation among wild-type lines for response to the drugs is difficult to establish. An interval mapping design was also used to map two or three QTL for each behavioral trait in a set of recombinant inbred lines derived from the laboratory stocks Oregon-R and 2b.