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Experimental test and refutation of a classic case of molecular adaptation in Drosophila melanogaster

Nature Ecology & Evolution volume 1, Article number: 0025 (2017) Cite this article

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Abstract

Identifying the genetic basis for adaptive differences between species requires explicit tests of historical hypotheses concerning the effects of past changes in gene sequence on molecular function, organismal phenotype and fitness. We address this challenge by combining ancestral protein reconstruction with biochemical experiments and physiological analysis of transgenic animals that carry ancestral genes. We tested a widely held hypothesis of molecular adaptation—that changes in the alcohol dehydrogenase protein (ADH) along the lineage leading to Drosophila melanogaster increased the catalytic activity of the enzyme and thereby contributed to the ethanol tolerance and adaptation of the species to its ethanol-rich ecological niche. Our experiments strongly refute the predictions of the adaptive ADH hypothesis and caution against accepting intuitively appealing accounts of historical molecular adaptation that are based on correlative evidence. The experimental strategy we employed can be used to decisively test other adaptive hypotheses and the claims they entail about past biological causality.

A central goal of molecular evolutionary biology is to identify the genes and biological mechanisms that mediated historical adaptation. Rigorously testing hypotheses in this area has been a major challenge. Many studies infer past selection from statistical signatures in genes that are involved in biological processes that might have suited species to their environments1–4. But sequence signatures of selection can be forged by chance or demographic processes and it is difficult to predict from sequence alone how genetic changes affect phenotypes and fitness5–8. Compelling evidence for molecular adaptation therefore requires formulating and testing explicit hypotheses about the causal links between specific evolutionary changes in gene sequence and the resulting changes in molecular function, organismal phenotype and fitness6–10. Advances in genetic mapping, experimental studies of molecular function and transgenic engineering have allowed hypotheses of molecular adaptation between recently diverged populations to be tested with increasing rigour11–16. But hypotheses about adaptive divergence between species or at higher taxonomic levels are explicitly historical, so testing them requires the effect of genetic changes that occurred on phenotype and fitness in specific evolutionary lineages from the distant past to be measured. Here we address this challenge by combining ancestral protein reconstruction17 with biochemical experiments and physiological analysis of transgenic animals that carry ancestral genes.

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Figure 1: Predictions of the classic hypothesis of ADH adaptive evolution.
Figure 2: Effects of ADH sequence divergence on the activity of purified enzymes.
Figure 3: Effects of ADH sequence divergence on ethanol catabolism and fitness in transgenic flies.
Figure 4: Sequence evolution on the phylogeny of D. melanogaster and closely related species.

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Acknowledgements

We thank L. Picton, K. O’Brien, K. Gordon and members of the C. Meiklejohn and K. Montooth laboratories for technical assistance. We thank D. Matute for providing polymorphism data for D. yakuba. We thank M. Kreitman, members of the J. Thornton laboratory and D. Anderson for comments and suggestions that enriched the project. The project was supported by a National Science Foundation (NSF) grant (DEB-1501877; J.W.T./M.A.S.), an NSF graduate research fellowship (M.A.S.), National Institutes of Health (NIH) grant (R01-GM104397; J.W.T.), NSF CAREER Award (1505247; K.L.M.) and an NIH training grant (T32-GM007197; M.A.S.). D.W.L. was supported by a Howard Hughes Medical Institute postdoctoral fellowship from the Life Sciences Research Foundation and an investigatorship to S. B. Carroll from the Howard Hughes Medical Institute.

Author information

Authors and Affiliations

  1. Department of Ecology and Evolution, University of Chicago, Chicago,, Illinois, USA

    Mohammad A. Siddiq & Joseph W. Thornton

  2. Laboratory of Cell & Molecular Biology, University of Wisconsin-Madison, Madison, Wisconsin, USA

    David W. Loehlin

  3. Howard Hughes Medical Institute, University of Wisconsin-Madison, Madison, Wisconsin, USA

    David W. Loehlin

  4. School of Biological Sciences, University of Nebraska, Lincoln, Nebraska, USA

    Kristi L. Montooth

  5. Department of Human Genetics, University of Chicago, Chicago, Illinois, USA

    Joseph W. Thornton

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  1. Mohammad A. Siddiq
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Contributions

M.A.S. and J.W.T. conceived the project. All authors participated in the experimental design. M.A.S. performed the phylogenetic and population genetic analyses. D.W.L. constructed the transgenic animals. M.A.S., D.W.L. and K.L.M. performed the functional experiments. All authors participated in data analysis and interpretation. M.A.S. and J.W.T. wrote the paper with contributions from D.W.L. and K.L.M.

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Correspondence to Joseph W. Thornton.

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Siddiq, M., Loehlin, D., Montooth, K. et al. Experimental test and refutation of a classic case of molecular adaptation in Drosophila melanogaster. Nat Ecol Evol 1, 0025 (2017). https://doi.org/10.1038/s41559-016-0025

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