Evidence that Environmental Heterogeneity Maintains a Detoxifying Enzyme Polymorphism in Drosophila melanogaster

Abstract

Environmental heterogeneity is thought to be an important process maintaining genetic variation in populations [1-4]: if alternative alleles are favored in different environments, a stable polymorphism can be maintained [1, 5, 6]. This situation has been hypothesized to occur in genes encoding multi-substrate enzymes [7], in which changes that increase activity with one substrate typically decrease activity with others [8-10], but examples of polymorphisms maintained by this mechanism are rare. Here, we present evidence that a polymorphism in an enzyme gene in Drosophila melanogaster is maintained by such a trade-off. The mitochondrially localized aldehyde dehydrogenase in D. melanogaster has two important functions: detoxifying acetaldehyde derived from dietary ethanol [11] and detoxifying larger aldehydes produced as byproducts of oxidative phosphorylation [12]. A derived variant of the enzyme, Leu479Phe, is present in moderate frequencies in most temperate populations but is rare in more ethanol-averse tropical populations. Using purified recombinant protein, we show that the Leu-Phe substitution increases turnover rate of acetaldehyde but decreases turnover rate of larger aldehydes. Furthermore, using transgenic fly lines, we show that the substitution increases lifetime fitness on medium supplemented with an ecologically relevant ethanol concentration but decreases fitness on medium lacking ethanol. The strong, opposing selection pressures, coupled with documented highly variable ethanol concentrations in breeding sites of temperate populations, implicate an essential role for environmental heterogeneity in maintaining the polymorphism.

Figure 1. Change in frequency of the Phe allele in experimental populations reared in the absence or presence of ethanol

Populations started with equal frequencies of Leu and Phe transgenic alleles (dashed line), and were genotyped after nine generations. Bars show ±1 binomial standard error. See also Figure S1.

Figure 2. Kinetic and structural properties of purified ALDH variants

(A) Turnover rates (± S.E.M.) with saturating levels of aldehydes of a range of molecular weights (1 mM substrate). The difference between Leu and Phe variants is statistically significant for each substrate (acetaldehyde, P < 0.01; butanal, P < 0.05; others, P < 0.001). (B) Turnover rates with two damaging aldehydes generated in mitochondria by lipid peroxidation (1 uM substrate). The difference between Leu and Phe variants is statistically significant for each (P < 0.002). (C) Superposition of Phe479 (red) on Leu479 (blue) in model structure of D. melanogaster ALDH. The catalytic residue, Cys322, is colored in yellow. Distances between Cys322 and Phe479 or Leu479 sidechains are shown to demonstrate the relative occupancy of the two sidechains within the substrate entry channel (SEC). (D) The Leu479Phe substitution constricts the SEC proximal to the substrate binding site, reducing the volume of the channel from 535ų to 456ų. The inner ring (red) represents the SEC diameter in ALDH^Phe479 and the outer ring (blue and green) represents the SEC diameter in ALDH^Leu479. See also Figures S2 and S3.

Figure 3. Survival under hyperoxia and acetaldehyde stress

Leu lines have significantly higher survival than Phe lines (P < 0.05) under hyperoxia, but significantly lower survival (P < 0.02) in the presence of acetaldehyde vapor. P-values are one-tailed.