Multiple mechanisms drive genomic adaptation to extreme O2 levels in Drosophila melanogaster

Abstract

To detect the genomic mechanisms underlying evolutionary dynamics of adaptation in sexually reproducing organisms, we analyze multigenerational whole genome sequences of Drosophila melanogaster adapting to extreme O₂ conditions over an experiment conducted for nearly two decades. We develop methods to analyze time-series genomics data and predict adaptive mechanisms. Here, we report a remarkable level of synchronicity in both hard and soft selective sweeps in replicate populations as well as the arrival of favorable de novo mutations that constitute a few asynchronized sweeps. We additionally make direct experimental observations of rare recombination events that combine multiple alleles on to a single, better-adapted haplotype. Based on the analyses of the genes in genomic intervals, we provide a deeper insight into the mechanisms of genome adaptation that allow complex organisms to survive harsh environments.

Fig. 1. Strong environmental selection pressure leading to the various alterations in the L- and H-population.

a Plot depicting oxygen level and the generations for L-population (blue line) and H-populations (orange line) (Source data are provided as Fig. 1a Source Data), b Estimated vs observed population size and error bands correspond to 95% confidence interval for the regression coefficient (Pearson’s R) (Source data are provided as Fig. 1b Source Data), c Estimated population size at different generations under selection pressure of L and H environments (Source data are provided as Fig. 1c Source Data), and d Principal Component Analysis (PCA) depicting PC1 and PC2 for the three replicates of L-, H-, and N-populations explains 45% of the variance (Supplementary Fig. 1a). The PCA was performed using only extant single nucleotide polymorphisms (SNPs) (Source data are provided as Fig. 1d Source Data). L-population, the population evolving in hypoxic environments; H-population, the population evolving in hyperoxic environments; N-population, the population maintained in normoxic environment.

Fig. 2. Deciphering the underlying mechanisms of selection using the Experimental Evolution Selection Analysis Pipeline (ESAP).

a ESAP takes a time-series interval of pooled frequencies from a genomic region and predicts the mechanism of the selection sweeps based on allele-frequency trajectories. b The characteristic allele-frequency trajectory for a hard-sweep simulation, when the favored mutation lies on a homogeneous background. The trajectories of all linked mutations on that haplotype converge to fixation. c The characteristic allele-frequency trajectory in simulations of a soft sweep due to standing variation. The favored mutation itself is fixed along with tightly linked mutations (black lines). However, the favored mutation is carried by multiple haplotypes, which drift at intermediate frequencies (orange lines). d Simulated instance of favored de novo mutation in a ‘late’ sweep. e Simulation of an ‘FM recombination’ that combines beneficial mutations onto a single haplotype. Starting at generation 80 from a cluster (haplotype) of fixed alleles, and tracing back in time, ESAP identifies two distinct clusters/haplotypes colored blue and orange (top panel). The blue and orange circles in the middle and lower panels provide the allele frequencies and genomic location of the mutations in these haplotypes at generation 1 (middle panel), and generation 80 (lower panel). Note the perfect separation of the two flanking haplotypes around ChrX:7,750,000 i.e., the recombination locus.

Fig. 3. Mechanisms of genetic adaptation utilized by Drosophila melanogaster in extreme O₂ environments.

a Manhattan plot showing five replicated sweeps in the L-population. Allele-frequency trajectories of an early hard sweep (region L_D) in the three replicates of populations evolving under low O₂. The x-axis is on a logarithmic scale and shows fixation by generation 34. b A replicated soft sweep due to standing variation in interval H_D. The favored allele was fixed at generation 61 while a second, tightly linked cluster (orange color) remains at intermediate frequency. c A de novo mutation in interval L_2A in one replicate of the L-population. d An FM-recombination in interval H_1B in one replicate of the H-population. Alleles that are fixated in generation 180 form two distinct clusters at generation 114 (orange and blue colors, respectively) that are spatially separated on chromosome X (position ChrX:7,750,000).

Fig. 4. Representative molecular functions of the candidate genes depicting overrepresentation of four major signaling pathways critical for regulating hypoxia tolerance.

a Rho guanyl-nucleotide exchange factor activity (pink area, p-value = 5.65E-06), b glutamate receptor activity (purple area, p-value = 2.31-06), c PI3K activity (green area, p-value = 2.24E-06) and d Vascular endothelial growth factor (VEGF) signaling (orange area, p-value = 4.45E-06). The p-value was calculated using Hypergeometric test and corrected using Benjamini & Hochberg False Discovery Rate (FDR). A p-value < 0.05 as significant level.