One of the challenges of evolutionary biology is understanding the genetic basis of adaptation. Many evolutionary changes occur with relatively little genetic change. For example, humans and chimpanzees differ greatly in anatomy, physiology, and behavior, yet they differ at only about 4% of their DNA sequences. Evolutionary biologists have long assumed that many anatomical differences result not from the evolution of new genes, but rather from relatively small DNA differences that alter the expression of existing genes. Recent research in evolutionary genetics has focused on how evolution occurs through alteration of gene expression.

An example of adaptation that has occurred through changes in regulatory sequences is seen in the evolution of pigmentation in Drosophila melanogaster fruit flies in Africa. Most fruit flies are light tan in color, but flies in some African populations have much darker abdomens. These darker flies usually occur in mountainous regions at higher elevations. Indeed, 59% of pigmentation variation among populations within sub-Saharan Africa can be explained by differences in elevation (Figure 18.19). Researchers have demonstrated that these differences are genetically determined and that natural selection has favored darker pigmentation at high elevations. High-elevation populations are exposed to lower temperatures, and the darker pigmentation is assumed to help flies absorb more solar radiation and better regulate their body temperature in these environments.

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How did flies at high elevations evolve darker color? Genetic studies indicate that the dark abdominal pigmentation seen in flies from these populations results from variation at or near a locus called ebony. The ebony locus encodes a multifunctional enzyme that produces a yellow exoskeleton; the absence of this enzyme produces a dark phenotype. Sequencing of the ebony locus of flies from light and dark populations found no differences in the coding region of the ebony gene. However, molecular analysis revealed a marked reduction in the amount of ebony mRNA in darker flies, suggesting that the difference in pigmentation is due not to mutations in the ebony gene itself, but rather to differences in its expression. Further investigation detected genetic differences within an enhancer that is about 3600 bp upstream of the ebony gene. Dark and light flies differed in over 120 nucleotides scattered over 2400 bp of the enhancer. By experimentally creating enhancers with different combinations of these mutations, researchers determined that five of the mutations are responsible for the majority of the difference in pigmentation.

These studies suggested that over time, high-elevation populations accumulated multiple mutations in the enhancer, which reduced the expression of the ebony locus and caused darker pigmentation. Further analysis suggested that these mutations were added sequentially. Some of the mutations are widespread throughout Africa; it is assumed that these existing mutations were favored by natural selection in high-elevation populations and increased in frequency because they helped the flies thermoregulate in colder environments. Other mutations that are seen only in the high-elevation populations probably arose as new mutations within those populations and were quickly favored by natural selection.