In their classic paper “Evolution at Two Levels in Humans and Chimpanzees,” published exactly 30 years ago, Mary-Claire King and Allan Wilson described the great similarity between many proteins of chimpanzees and humans . They concluded that the small degree of molecular divergence observed could not account for the anatomical or behavioral differences between chimps and humans. Rather, they proposed that evolutionary changes in anatomy and way of life are more often based on changes in the mechanisms controlling the expression of genes than on sequence changes in proteins.
This article was a milestone in three respects. First, because it was the first comparison of a large set of proteins between closely related species, it may be considered one of the first contributions to “comparative genomics” (although no such discipline existed for another two decades). Second, because it extrapolated from molecular data to make inferences about the evolution of form, it may also be considered a pioneering study in evolutionary developmental biology. And third, its focus on the question of human evolution and human capabilities, relative to our closest living relative, marked the beginning of the quest to understand the genetic basis of the origins of human traits. Like much of Wilson and his colleagues' body of work, this contribution had a great influence on paleoanthropologists as well as molecular biologists.
The 30th anniversary of this landmark article arrives at a moment when comparative genomics, evolutionary developmental biology, and evolutionary genetics are pouring forth unprecedented amounts of new data, and the entire chimpanzee genome is available for study. It is therefore an opportune time to examine what has been and is being revealed about the relationship between evolution at the two levels of molecules and organisms, and to assess the status of King and Wilson's hypothesis concerning the predominant role of regulatory mutations in organismal evolution.
King and Wilson used the phrase “ways of life” to include both physiology and behavior (M.-C. King, personal communication) and proposed that the evolution of both anatomy and ways of life was governed by regulatory changes in the expression of genes. From the outset of this review, I make the sharp distinction between the evolution of anatomy and the evolution of physiology. Changing the size, shape, number, or color patterns of physical traits is fundamentally different from changing the chemistry of physiological processes. There is ample evidence from studies of the evolution of proteins directly involved in animal vision , respiration , digestive metabolism , and host defense  that the evolution of coding sequences plays a key role in some (but not all) important physiological differences between species. In contrast, the relative contribution of coding or regulatory sequence evolution to the evolution of anatomy stands as the more open question, and will be my primary focus.
The amount of direct evidence currently in hand is modest, and includes examples of both the evolution of coding and of non-coding, regulatory sequences contributing to morphological evolution. However, I will develop the argument, on the basis of theoretical considerations and a rapidly expanding body of empirical studies, that regulatory sequence evolution must be the major contributor to the evolution of form.
This conclusion poses particular challenges to comparative genomics. While we are often able to infer coding sequence function from primary sequences, we are generally unable to decipher functional properties from mere inspection of non-coding sequences. This has led to a bias in comparative genomics and evolutionary genetics toward the analysis and reporting of readily detectable events in coding regions, such as gene duplications and protein sequence evolution, while non-coding, regulatory sequences are often ignored. However, approximately two-thirds of all sequences under purifying selection in our genome are non-coding . One consequence of the underconsideration of non-coding, regulatory sequences is unrealistic expectations about what can currently be learned about the genetic bases of morphological diversity from comparisons of genome sequences alone. The visible diversity of any group is not reflected by the most visible components of gene diversity—that is, the diversity of gene number or of coding sequences. In order to understand the evolution of anatomy, we have to study and understand regulatory sequences, as well as the proteins that connect them into the regulatory circuits that govern development. I will begin with some historical and theoretical considerations about regulatory and coding sequence evolution, then delve into the insights offered by specific experimental models of anatomical evolution, and finally, I will revisit King and Wilson's original focus and discuss how our emerging knowledge of the evolution of form bears on current efforts to understand human evolution.