Extreme QTL mapping enables the identification of novel zinc toxicity response loci in Drosophila melanogaster.

Heavy metals are a widespread environmental contaminant, and even low levels of some metals can disrupt cellular processes and result in DNA damage. However, the consequences of metal exposure are variable among individuals, with susceptibility to metal toxicity representing a complex trait influenced by genetic and non-genetic factors. To uncover toxicity response genes, and better understand responses to metal toxicity, we sought to dissect resistance to zinc, a metal required for normal cellular function, which can be toxic at high doses. To facilitate efficient, powerful discovery of Quantitative Trait Loci (QTL), we employed extreme, or X-QTL mapping, leveraging a multiparental, recombinant Drosophila melanogaster population. Our approach involved bulk selection of zinc-resistant females, sequencing several replicate pools of selected and control animals and identified QTL as genomic positions showing consistent allele frequency shifts between treatments. We successfully identified 7 regions segregating for resistance/susceptibility alleles and implicated several strong candidate genes. Phenotypic characterization of populations derived from selected or control animals revealed that our selection procedure resulted in greater egg-to-adult emergence and a reduced developmental delay on zinc media. We subsequently measured emergence and development time for a series of midgut-specific RNAi gene knockdowns and genetic controls raised in both zinc-supplemented and normal media. This identified 10 genes with significant genotype-by-treatment effects, including pHCl-2, which encodes a zinc sensor protein. Our work highlights recognized and novel contributors to zinc toxicity resistance in flies and provides a pathway to a broader understanding of the biological impact of metal toxicity.