Try to outrun a horse and you’ll likely lose. Despite the persistent myth that our species evolved to chase down prey on the savanna, horses will generally beat even the most elite human endurance athletes in contests of speed and distance. New research helps explain why and how horses have an athletic advantage.
A study published March 27 in the journal Science finds that a unique and ancient spate of genetic mutations was key to the evolution of horses’ exceptional aerobic fitness. Together, these changes enable horse muscles to use oxygen quickly and efficiently, without incurring the cellular damage normally associated with burning through lots of fuel.
“It really lets horses have their cake and eat it too,” Gianni Castiglione, a study co-author and a biologist at Vanderbilt University, tells Popular Science. The mutations partially account for why horses are able to maintain such a high proportion of muscle relative to their body mass, why they can sustain such a high concentration of mitochondria in their muscle cells, and why their maximum ability to uptake and use oxygen (or their VO2 max) is more than double that of a human athlete.
The gene and resulting protein at the center of the adaptation isn’t just for horses– it also has serious clinical implications for health research. It’s possible that, by uncovering the mystery of horse athleticism, scientists could find new treatments for neurodegenerative diseases in humans like Alzheimer’s or even therapies to stall the declines of normal aging.
A first of its kind mutation in mammals
Castiglione and his team began their quest with a broad survey of genes across the tree of animal life, looking for how genetic sequences with known importance in people differ or remain the same in other species. Unexpectedly, they homed in on a specific equine alteration that helps explain how horses (along with zebras and donkeys) achieve their propulsive power. “We found this entire phenomenon in horses–something we would have never predicted,” Castiglione says.
Adding to the intrigue, the change in horses is closely related to one also found in birds, says Elia Duh, a study co-author and an ophthalmologist and biomedical scientist at Johns Hopkins University. Castiglione and Duh published previous research in 2020 on that avian discovery, and believe the change is a major part of why birds are able to fly, despite how energetically taxing all of that flapping is.
“It implied convergent evolution,” Castiglione says. But once the scientists dug deeper, they found that the genetic shift in horses is far more complicated than the bird mutation. Horses, it turns out, had to clear many more evolutionary hurdles to make it to their final, fit form. One of these hurdles is so rare, it’s a type of mutation only previously seen in a subset of viruses.
The protein factor
To understand how horses found their athletic stride, it’s first important to understand two proteins: NRF2 and KEAP1. NRF2 is basically identical across almost all vertebrates. It has antioxidant effects in the body, neutralizing damaging molecules. It also plays a role in the production of the molecule adenosine triphosphate (ATP), which is the energetic currency of cells. Mitochondria AKA the “powerhouse of the cell” burn ATP as fuel. The more ATP available, the more work cells can do.
Though NRF2 performs some very important functions, it can also be deadly if it runs amok. In many vertebrates, overactive NRF2 triggers big problems like tumor growth. “It’s very much a Goldilocks situation,” says Castiglione. Animals “have to tightly regulate NRF2 and only turn it on when they need it.”
[ Related: Why studying horses could help humans stay healthy, too. ]
That’s where KEAP1 comes in. It keeps NRF2 in check by glomming onto NRF2 and clogging up its active site under normal cellular conditions. When enough oxidizing molecules, also called free radicals, show up as an unavoidable byproduct of cells using energy, KEAP1 gets deactivated. This frees NRF2 to fight off the free radicals and minimize cellular harm.
In birds, changes to the gene that codes for KEAP1 render the protein non-functional. Avians developed strategies to compensate for some of the consequences of having unregulated, constantly active NRF2 proteins–while still reaping the reward of enough energy to fly. “There’s definitely a balance, and–for different organisms–it’s a matter of what’s the optimal balance,” Duh says.
Horses harness the power of ‘recoding’
In horses and their close relatives, things are a little more convoluted. The study authors found a single nucleotide change early on in the gene that codes for KEAP1, which would normally halt protein production. However, through protein sequencing, cell culture experiments, and comparisons with mouse and human cells, Duh and Castiglione discovered that horses don’t have a non-functional, stunted protein. Instead, the equine version of KEAP1 is even more sensitive to stress, and more responsive to free radicals than the kind that exists in other vertebrates.
Horses do this through a phenomenon called recoding. During recoding, a gene sequence normally read as a stop sign is translated into an amino acid which allows the rest of the protein to be built. A suite of complementary genetic changes had to happen to enable the recoding, but it’s not clear if those came before or after the KEAP1 gene mutation itself. Either way, the resulting horse KEAP1 protein is just different enough to offer benefits, without any obvious, major downsides.
This particular type of recoding has only previously been documented in viruses that infect bacteria–or bacteriophages. Finding the process in a vertebrate mammal reveals a whole new realm of possible genetic weirdness lurking in complex organisms. “It just speaks to the uniqueness of horses,” says Duh.
[ Related: Horses once had multiple hoofed toes. ]
The sheer number of DNA shifts that must have occurred in rapid succession to create this synchronous outcome also reflects the strong pressure horses were likely under to become fast and tireless. The first horse-like animals were about the size of dogs and faced several predators in their open grassland habitat. To avoid being eaten to extinction, they adapted. It was an “intense evolutionary crucible” and ancestral horses had to innovate to survive, suggests Castiglione.
It’s not clear exactly when in horse history these changes happened. It could have been just before the origins of the genus Equus between 4 and 4.5 million years ago. Or, it may have been as distant as 55 million years ago, just after the common ancestors of contemporary rhinoceros and horses split. The genetic record is too incomplete to draw detailed conclusions about the timeline, Castiglione notes. “We can only infer what happened once upon a time– unless you have a time machine. That’s always a limitation with evolutionary work.”
Future applications in humans
Despite the unanswered evolutionary questions, Castiglione and Duh believe that their findings are firm enough to spur biomedical advances. KEAP1 and NRF2 are critically important proteins in many human diseases and in aging. Knowing that a single DNA nucleotide and amino acid swap can significantly change how the protein complex functions could be useful for addressing some of the malfunctions that pop up.
Then there’s the fact that horses evolved a workaround to avoid getting stuck with a shoddy KEAP1 protein, despite a mutation that should have halted the production line. “In 10 to 15 percent of human diseases, you have a premature stop codon,” says Castiglione. If horse ancestors found a way to still produce a functional protein, perhaps a similar, equine-inspired gene therapy could be developed for humans too.
Yet for those future possibilities to materialize, there needs to be continued investment in exploratory and fundamental research, says Castiglione. “All of this work was funded by the [National Institutes of Health],” he tells Popular Science. “It’s because of their willingness to fund basic research that now we’ve found things of clinical value that we would have never been able to predict.” Under the current administration, NIH is facing major funding cuts.
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