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Snakes used to wander the Earth on legs about 150 million years ago, before they shifted from strut to slither.

Now, two scientists have pinpointed the genetic process that caused snakes to lose their legs. What's more, they say the "molecular machinery" for leg development still persists in snakes after these millions of years -- it's simply switched off.

Some snakes, such as pythons, retain tiny vestiges of legs in the form of two small bumps on either side of their pelvis. The scientists at the University of Florida who conducted the snake-leg research published Thursday in Current Biology focused on pythons for the study.

As co-author and molecular genetics professor Martin Cohn tells The Two-Way, a gene known as Sonic hedgehog is responsible for limb development. (Yes, this gene was named by enthusiasts of the video game.)

The Florida researchers found that the Sonic hedgehog gene flickers on very briefly in python embryos less than 24 hours old. That genetic signal hadn't previously been detected in pythons.

UF Health researchers uncover how snakes lost their legs

Cohn says the snakes' Sonic hedgehog turns on as in any vertebrate with limbs, but then switches off. "We really wanted to drill down," he says, "to the exact nature of the reason for that in the genome."

He explains that in limbed vertebrates, Sonic hedgehog controls the development of many organs, not just arms and legs. DNA regulatory sequences called enhancers control the way these genes switch on and off for specific body parts. Without an enhancer for limbs, Sonic hedgehog could not make them fully develop. The researchers found that several sections of DNA in that enhancer were deleted at some point during python evolution.

The same sections were deleted in the DNA sequences of boa constrictors, Cohn says. And the results were even more pronounced in snakes that no longer have any vestiges of legs at all, such as cobras and vipers. Their Sonic hedgehog limb enhancer was "almost undetectable," he says. "So this tells us that there has been sequential degradation of Sonic hedgehog in the limbs in snakes."

To recap, the study says these mutations of the Sonic hedgehog limb enhancer are responsible for arrested limb development in pythons.

The research then took an even more surprising turn when the scientists studied the activity of another gene, called HOXD, that's involved in making fingers and toes. While studying python embryos, "we saw a pattern of activity of this HOXD gene that was characteristic of a foot, which is a really strange result, because pythons don't build feet," Cohn says.

And then, Cohn says, he and fellow researcher Francisca Leal found this: Embryonic pythons "make a pre-cartilaginous model of all of the skeletal elements of the limb. ... They start forming a complete leg all the way down to the foot."

"It was absolutely mind-blowing because this was like development of a cryptic leg skeleton in an embryo," he says, adding that the mutated Sonic hedgehog limb enhancer ultimately doesn't allow this cryptic limb to grow.

Cohn says this points to an interesting implication about the way snakes have evolved over millions of years:

"There are a number of fossil snakes with legs. Paleontologists have argued that some of these ancient snakes retained the legs of their ancestors, whereas others species evolved after legs were lost, and, thus, they may have reacquired legs."

That could be because pythons have retained much of the molecular machinery for making limbs, according to Cohn.

"It's fascinating -- how do you re-evolve a structure that was lost? It's pretty uncommon," he says.

"But what these results tell us is that maybe that wasn't as challenging a process as it might seem. They didn't have to reinvent limbs from scratch. There's already a remarkable degree of limb development that happens in the embryo and if you just retained those embryonic structures and allowed them to complete skeletal development, you have legs."

Science

Scientists in Florida say they've pinpointed a genetic process that caused snakes to lose their legs and have found that embryonic pythons still form "cryptic leg skeletons," millions of years later.

How snakes lost their legs

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