A genome-wide, fine-scale map of natural pigmentation variation in Drosophila melanogaster

Abstract

Various approaches can be applied to uncover the genetic basis of natural phenotypic variation, each with their specific strengths and limitations. Here, we use a replicated genome-wide association approach (Pool-GWAS) to fine-scale map genomic regions contributing to natural variation in female abdominal pigmentation in Drosophila melanogaster, a trait that is highly variable in natural populations and highly heritable in the laboratory. We examined abdominal pigmentation phenotypes in approximately 8000 female European D. melanogaster, isolating 1000 individuals with extreme phenotypes. We then used whole-genome Illumina sequencing to identify single nucleotide polymorphisms (SNPs) segregating in our sample, and tested these for associations with pigmentation by contrasting allele frequencies between replicate pools of light and dark individuals. We identify two small regions near the pigmentation genes tan and bric-à-brac 1, both corresponding to known cis-regulatory regions, which contain SNPs showing significant associations with pigmentation variation. While the Pool-GWAS approach suffers some limitations, its cost advantage facilitates replication and it can be applied to any non-model system with an available reference genome.

Figure 1. Overview of the experimental design.

Wild D. melanogaster flies were collected from Vienna, Austria, and Bolzano, Italy, brought into a controlled environment in the laboratory, and treated as shown in the figure. The same procedure was used for all five replicates, with each replicate resulting in 1,500 females for phenotyping, and with 100 light and dark flies from each replicate sequenced.

Figure 2. Genome-wide association study of female abdominal pigmentation.

A) Manhattan plot for abdominal pigmentation in the full data set (including all five replicates). The–log₁₀ p-values are plotted against the position on each chromosome. The horizontal dashed line indicates the genome-wide significance threshold at an FDR of 0.05. The red bars indicate candidate genes previously shown to affect pigmentation. B) Detailed view of the tan (left), bab1 (middle) and ebony (right) regions. The positions on the x-axis are indicated in kb. The red bars indicate regulatory regions previously shown to affect pigmentation. C) Detailed view of top ranked polymorphisms located within or close to the three pigmentation genes. For every gene, the most significant SNPs fall in regulatory regions.