A leg growing out of your mouth. Extra legs pushing out of your side. Walking parts where your swimming bits should be. If you’re a tiny crustacean in Nipam Patel’s lab, chances are good you’re not quite right—and that’s just the way these UC Berkley geneticists like it. By inducing birth defects in arthropods called parhyale, Patel’s team makes “monsters” that deliver a one-two punch, offering insights into the mechanics of evolution, and into ways we could treat (or even prevent) human birth defects and disease in the future.
The questions that drive Patel’s lab are deceptively simple, and brain-crushingly profound: How did Earth’s organisms become different from one another? How is an embryo “programmed” to know what it should look like? How might changes to that programming have advanced evolution itself? This frustrated, flailing parhyale–which, thanks to Patel’s crew, was born with almost perfect walking appendages where its swimming appendages should be–is helping to revolutionize how we think about all of it.
Welcome to the world of Hox genes, a roughly 600-million-year-old “toolkit” that controls how body plans–the head-to-tail layout of our symmetrical, physical selves–develop. Once thought to exist only in flies, Hox genes rocked biology in the mid-’80s when it was discovered that they were in every single animal on Earth. And while the number of Hox genes tends to vary according to how complex you are (insects have 8; humans have 39), the genes themselves have changed so little in millions of years that they’re what’s called “highly conserved” across species.
In labs, that means flies function surprisingly well when one of their Hox genes is swapped for the corresponding chicken Hox gene. From an evolutionary perspective, it means earthworms, humpback whales, butterflies, and humans are all just variations on a theme. “Despite the fact that we don’t think of ourselves as looking anything like a fly,” says Patel, “our development basically uses the same genes.”
Hox genes are “master instructors”–each oversees development in a different region of the body (head, thorax, abdomen), turning other genes on and off to ensure you grow the right form for your species. “In the field in general,” says Patel, “I think we’ve increasingly convinced people that single genes can have big roles in evolution,” but his team hunts proof, examples of how small tweaks to the Hox toolkit may have given rise to Earth’s massive species diversity.
Patel’s lab focuses on crustaceans, a massively diverse group (67,000 known species) that feature what he describes as “the Swiss Army-knife approach to a body plan, with lots of different legs doing lots of different things.” Within this group, parhyale is a dreamboat model organism: easy to breed, easy to harvest embryos from, and “because of the way the embryo develops,” says postdoc Arnaud Martin, “we can make one half of it mutant and the other half normal, giving us a built-in control.”
Patel, Martin, and grad students Erin Jarvis and Heather Bruce use two techniques to tweak embryos’ Hox genes: RNA Interference, which “knocks-down” genes by inhibiting their expression, and the ridiculously cutting-edge CRISPR/Cas9 genome-editing system, which “knocks-out” genes with a level of precision that was unimaginable even two years ago. The resulting mutants are dramatic enough to earn nicknames in the lab–“Shiva” is loaded with extra legs on one side, while “Foot in Mouth” has a gangly limb poking out of a maw that should house a small, demure filtering appendage–and they teach researchers more about how development processes (and Hox genes themselves) work. But they’re not all Patel’s team is looking for.
“Those extreme phenotypes are going to die,” says Patel. “They’re not the stuff of evolution. But the beauty,” he adds, proverbial twinkle in his eye, “is that we also get a range of very weak phenotypes that could definitely be the stuff of evolution.”
The lab’s weak phenotypes are a lot less dramatic (a slightly wonky head bristle instead of a full transformation to head-leg, for example), but they bring to life “the kind of stuff you can imagine evolution actually doing”–and they help Patel’s researchers investigate how evolution gets it done. One mechanic they think evolution employs is to slightly shift the borders of regions controlled by various Hox genes, and the group is already known for one piece of evidence connecting such shifts to species diversity. By altering the anterior (head-direction) “expression boundary” of a Hox gene called Ubx, they created parhyale with varying numbers and arrangements of feeding appendages, patterns that replicate those seen in thousands of other crustacean species.
“In order for that to be a true source of evolutionary change, though” says Jarvis, “it can’t just be skin-deep; transformations have to extend to the musculature and nervous systems.” So the lab (and Jarvis in particular) also peer below, and get their answers in neon lights. In one mutant, a fluorescent green stain reveals the totally functional musculature of a near-perfect walking appendage—a limb that would have been a swimming appendage had Jarvis not knocked down a Hox gene called Abd-B.
“Basically, we take a sledgehammer approach in the lab,” says Martin. “But versions of what we do here might be what happens in nature to create diversity.”
Patel’s team plans to publicize two new pieces of supporting evidence soon, but already another question looms large over future research: What actually causes changes to the way Hox genes express themselves? In the lab, the culprits are clear, but what mechanism is at work in the evolutionary wild?
As for how this kind of research might apply outside the lab, Patel admits he’s not eager for us to head down the “slippery slope” of human-genome editing, but he’s also quick to point out that understanding what Hox genes do during normal development is critical to understanding how and why they sometimes go wrong. In humans, inappropriate Hox gene expression causes a wide range of birth defects and disease, including several forms of childhood leukemia.
“The ability clinicians have now to find a mutation in humans and say ‘this is the causal gene,’ it’s amazing.” Patel says. “But even when you make the discoveries in humans, you still can’t study them directly that way. To understand it, to prove it, to address it, you have to go to a model system like parhyale.”