Athens, Ga. – When researchers at the University of Georgia examined the genome of several different snake species, they found something surprising. Embedded in reptiles’ genetic code was DNA that, in most animals, controls the development and growth of limbs—a strange feature for creatures that are famous for their long, legless bodies and distinctive slither.

Now, they’ve found an explanation. In a paper published today in the journal Developmental Cell, the scientists show that the same genetic tools responsible for limb development also control the formation of external genitalia, and that may help explain why snakes have held on to this limb circuitry through the ages.

Snakes weren’t always legless; they evolved the loss of limbs over 100 million years ago, said Douglas Menke, an assistant professor of genetics in UGA’s Franklin College of Arts and Sciences and senior author of the paper.

“There have been many millions of snake generations since they evolved a legless body, and we would generally expect the DNA associated with limb development to fade away or mutate to do another job, but that doesn’t seem to have happened,” he said. “Naturally, we wanted to know why snakes had retained DNA that they don’t appear to need.”

In their experiments, Menke and postdoctoral researcher Carlos Infante examined specific regions of noncoding DNA known as enhancers—a kind of switch that controls the expression of genes, telling them when to turn on or off during embryonic development.

The researchers followed patterns of enhancer activity in embryonic limbs and genitalia of mice and lizards. This revealed that many of the same enhancers are activated during the formation of these different appendages in both species.

They engineered mice that lack one of these limb-genital enhancers and found defects in the legs and genitalia of the resulting mice. The snake version of this enhancer, however, only functions during development of genitalia.

“What this means is that much of the genetic circuitry that controls the development of limbs is also important for the formation of genitalia,” Menke said. “And we think that’s why snakes still have the genetic blueprints for limb development in their genome.”

It is generally accepted among evolutionary biologists that limbs evolved from fins, but the phallus—external genitalia that includes the penis and clitoris—is thought to be a much more recent development, he said. And there is evidence to suggest that the genes initially used to grow limbs were later co-opted for the development of a phallus.

“We’re only just beginning to understand the various roles of many of these enhancers,” Menke said. “But what we generally refer to as ‘limb enhancers’ should probably be more broadly categorized as ‘appendage enhancers,’ because they clearly perform more than one job.”

The research team had access to the genomes of three snake species for this study: boa constrictor, Burmese python and king cobra. Comparative genomics research like this has only recently been made possible as the genome sequences of snakes and other species have become available.

In future experiments, Menke will investigate the extent to which noncoding DNA influences the formation of different genital shapes observed in nature.

This work was supported by the National Science Foundation, the National Institutes of Health under grant number HD081034 and the University of Georgia.

The study is available at http://www.cell.com/developmental-cell/abstract/S1534-5807(15)00583-3.