I never could understand how monkeys keep their composure with swaths of mosquitos buzzing, bumping, and biting them to no end. Don’t they get itchy? Have they not heard about mosquito-borne viruses like dengue?
Pronounced DEN-gay or DEN-gee, the dengue virus infects 100 million people globally, killing 25,000 each year. After infecting a host, dengue can spread to other individuals or species through mosquito bites. In fact, dengue is “the most important mosquito-borne viral disease with epidemic potential in the world,” according to a 2014 report by the World Health Organization. Until now, the biology behind dengue’s surprisingly rapid transmission has baffled scientists hoping to develop a vaccine.
“A good animal model for infection doesn’t really exist right now,” said Alex Stabell, a researcher and medical student at the University of Colorado. In order to study the virus and create a vaccine, researchers typically need a living animal model. These can be primates, but small rodents are generally preferred. Despite being able to infect many mammals, researchers have yet to find any species that display symptoms like humans do, making it difficult to truly understand the virus. This (literal) life-or-death difference between species has largely been a mystery—but researchers are slowly cracking the code.
In a study published in the journal eLife, lead author Stabell and his colleagues focus on a protein called STING. STING is involved in the immune response to viruses. It can signal to the cell in such a way that the cell’s immune response is turned on. However, like a chain-link fence guarding hosts against viral infiltration, STING is only useful if intact. Stabell and his team demonstrate that dengue can recognize two specific links in that chain-link fence and—snip—cut it into two pieces. The links in the fence are the sequence of amino acids. “By knowing that sequence, we can search and find other animals that would either be susceptible or resistant,” said Stabell.
At the start of his PhD, Stabell was interested in a broader question: understanding how viruses influence the evolution of primates. Like a pesky mosquito in his ear, reports of dengue’s interaction with STING commanded his attention. The sequence of amino acids—the links in the chain—varies only slightly between species. Of 379 links in the human STING sequence, chimpanzees share all but three. In humans, previous studies showed that the dengue virus deactivates STING, allowing it to replicate rapidly and cause disease. But chimpanzees show no such symptoms. So how could it be that dengue behaves so differently in chimpanzees, which have a STING defense nearly identical to ours?
Give scientists an impactful problem that doubles as a puzzle, and you’re bound to pique their interest. Over the course of a few years, Stabell and other scientists working with him developed a platform to fundamentally understand how dengue and STING interact.
With the platform in place, Stabell infected cells with dengue virus and evaluated how different animal STING proteins offer protection against the virus. In addition to human STING, the team isolated the protein from non-human primates: chimpanzee, marmoset, and macaque. As expected, they found that dengue virus infection led to cleavage of human STING, but the virus left the other primate STING proteins surprisingly intact.
Recall that human and chimpanzee STING proteins differ in very few regions. One, the 78th link in the chain, stood out to Stabell. Previous reports estimated that STING cleavage occurred in this vicinity. Interestingly, where human STING has the amino acids arginine and glycine—represented as R-G—at positions 78 and 79, chimps have tryptophan and glycine, or W-G. To evaluate the importance of this segment, Stabell changed the 78th amino acid of human STING to match that of chimpanzees: W-G instead of R-G. Shockingly, this small change made the modified human STING entirely resistant to cleavage. Next, Stabell changed the chimpanzee STING from W-G to R-G at this position and found that it became newly susceptible to cleavage.
“You can make any STING protein cleavable or cleavage-resistant just by modifying one of two amino acids,” said Stabell. “That was the most surprising. And it’s all based on just this one little difference.”
The same result was shown for marmosets and macaques, whose unmodified STINGs have Q-G and R-D, respectively, at 78/79. This indicates that dengue virus can only deactivate STING proteins with the appropriate R-G motif. Without this motif, the protein chain stands intact, guarding the host from disease. We humans are out of luck and justified in our mosquito annoyance.
These findings may have immense bearing on the future of dengue studies. “Right now [scientists] use mice, but they have to eliminate most of their immune system for the virus to replicate effectively,” said Stabell. “Identifying certain things like the two amino acid motif…is one of probably many steps to making an animal model more human-like.”
Rather than rely on immune-compromised mice to model viral infection, Stabell envisions using the critical motif to select a model organism rationally. The team scoured GenBank, a database of protein and DNA sequences, for other animals with the R-G motif. They identify three small rodent species already used in other animal models—chinchillas, desert woodrats, and naked mole rats—as candidates.
Stabell suggested that future work may not be limited to just dengue. Other viruses in the dengue virus family, flaviviruses, can cleave immune-related proteins in a similar way. “There is some evidence that Hepatitis C virus…targets a protein that does essentially the same thing as STING does,” said Stabell. “There’s likely a commonality in all the flaviviruses that they target these innate immune proteins.”
Asked about how this multi-year project changed his perspective, Stabell responded that he’s in awe of dengue’s adapting to infect different hosts differently. “It gave me a better appreciation for the diversity of species and how elegant the viruses are that infect us all,” said Stabell. “That is one of the greatest mysteries and the thing that I will continue to be interested in forever.”
An estimated 3.9 billion humans risk infection from this easily transmissible virus. Thanks to work like Stabell’s, we have a better understanding of dengue and why most primate species are untroubled by some mosquito-borne diseases. Enlightening, surely, but hopefully we can soon decode their indifference to mosquito-borne ear-buzzing.
By Max Levy