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RBBP6 may hold the key
SAN FRANCISCO—Currently no drugs are available to treat Ebola, Dengue or Zika viruses, but new research from the Gladstone Institutes and the University of California, San Francisco (UCSF) may change that. Scientists have identified key ways the viruses hijack the body’s cells, and have found at least one potential drug that can disrupt this process in human cells. They have also discovered how the Zika virus may cause microcephaly in infants.
Published in back-to-back papers in the Dec. 13, 2018, issue of Cell, the researchers employed a technique called protein-protein interaction mapping to probe the virus trio. The scientists use these comprehensive maps to target interactions in order to try and kill infections.
“We’ve employed our systematic protein-protein interaction strategy on Ebola, Dengue and Zika to get a better sense of how these three very problematic viruses hijack, rewire and infect human cells,” says the leader of both studies, Dr. Nevan Krogan, who is a senior investigator at the Gladstone Institutes, director of QBI at UCSF and a professor of cellular and molecular pharmacology at UCSF. “To me, what’s most interesting is when we see the same human machinery being hijacked by seemingly very different viruses and different pathogenic proteins.”
The group’s first study, conducted with Dr. Christopher Basler at Georgia State University, identified 194 virus-human interactions involving six Ebola proteins. The scientists narrowed their focus to one point of contact between an Ebola protein called VP30 and a human protein called RBBP6. The interaction first caught their attention because it was such a strong one, but RBBP6 ended up being a particularly intriguing protein because it has emerged in other virus-protein interaction maps, leading Krogan to believe it plays an important role in the immune system.
It turns out that RBBP6 mimics another Ebola protein called NP. VP30 and NP need to bind to one another in order for the Ebola virus to replicate. However, the human protein RBBP6 interrupts this process by attaching to VP30 instead. So, by blocking the connection between the two Ebola proteins, RBBP6 effectively stops the virus from replicating. To their surprise, the researchers didn’t discover a way the virus attacks the host, but a way for the host to fend off the virus.
“We identified a relatively small domain in the human protein RBBP6 that is very similar to a region in the Ebola protein NP. We showed it’s the exact region in RBBP6 that binds to VP30. The interaction between NP and VP30 is required for Ebola infection, so RBBP6 is essentially competing with NP to bind to VP30,” points out Krogan.
“This mimicry is interesting because it’s quite unusual,” Krogan continues. “Evolutionarily, viruses evolve much more quickly than humans, so they use mimicry to develop resistance against the host. As a result, we normally see the virus mimicking a human protein for its own benefit. In this case, we found the opposite, because the host is using mimicry to imitate a viral protein and have an effect on the virus.”
“It appears our body has a natural way to fight off Ebola infection, and the virus hasn’t gotten around it,” noted Dr. Jyoti Batra, one of the first authors of the paper and a postdoctoral scholar at Gladstone, formerly in Basler’s laboratory at Georgia State University. “Keep in mind, we still don’t have great mechanisms to fight off Ebola, but without this protection the virus would be even deadlier.”
To test this theory, Batra worked with the study’s other first author, Dr. Judd Hultquist, who conducted the research as a postdoctoral scholar at Gladstone and UCSF and is now an assistant professor at Northwestern University Feinberg School of Medicine. Together, they engineered human cells to either have no RBBP6, or to have much higher levels than normal. Then they infected the cells with Ebola virus. In cells with no protective protein, infection rates went up fivefold. However, the cells with extra RBBP6 strongly prevented Ebola infection.
“We identified a 23-amino-acid peptide that, once we get it into cells, kills the Ebola virus. Using a peptide as a biologic drug has therapeutic value, but it also comes with challenges (not only to get it into the cells but also to ensure it doesn’t then get degraded),” says Krogan. “We also explored another option that may be more effective, based on the region of VP30 we identified to have clear therapeutic value.
“We want to look at the structure of VP30 that binds to RBBP6, and then computationally design a molecule or compound to mimic the binding location. We are currently working with structural computational biologists at UCSF who look at structures and can predict certain compounds to which they could bind.”
In the second paper, Krogan’s laboratory worked with researchers at Icahn School of Medicine at Mount Sinai and Baylor College of Medicine. The scientists hypothesized that if Dengue and Zika interact with human proteins in similar ways, targeting those protein interactions could be the best way to fight the infections. They also mapped the interactions between the Dengue virus and mosquito proteins to compare it to the human-virus protein maps.
“The virus replicates essentially in the same way in both human and mosquito cells,” mentioned Dr. Priya Shah, an assistant professor of chemical engineering and microbiology and molecular genetics at UC Davis who conducted the research as a postdoctoral scholar at UCSF. “So, if we can home in on the shared parts of these cells that are exploited by the virus, we could identify a potentially powerful therapeutic target.”
Comparing three interaction maps (Dengue-human, Zika-human and Dengue-mosquito), the scientists identified one interaction that occurred in both viruses and both host species: the viral protein NS4A and the host protein SEC61. SEC61 is known to play a critical role in some forms of cancer, and Krogan’s colleague at UCSF had developed compounds targeting these proteins as potential anti-cancer drugs.
“We found two compounds (CT8 and PS0361), both of which are drugs developed by Jack Taunton at UCSF. Taunton has shown these compounds bind to and inhibit the function of SEC61, which is a key protein involved in a crucial pathway. This pathway gets perturbed in different cancers and is also hijacked by the viruses to benefit infection,” explains Krogan. “Taunton has an ongoing clinical trial using these drugs as anticancer therapies. In our study, we found the same compounds also effectively wiped out both the Dengue and Zika infections when added to human and mosquito cells.”
“Now we need to tweak the molecule to optimize its safety and efficacy before it can be tested in patients,” said Taunton, a professor in the Department of Cellular and Molecular Pharmacology at UCSF.
Although Dengue and Zika are similar, only Zika causes microcephaly. Krogan’s team looked for examples where Zika proteins interacted with human proteins, while Dengue proteins did not. One interaction that stood out was between the Zika protein NS4A and the human protein ANKLE2, which is important for brain development. Mutations in ANKLE2 have previously been linked to hereditary microcephaly.
Scientists found that the Zika protein appears to inhibit the function of ANKLE2, impairing brain development and leading to microcephaly. The researchers plan to use this knowledge to start developing ways to target ANKLE2 to prevent Zika-related microcephaly.
“We determined that NS4A is inhibiting ANKLE2’s function, but we need to carry out more mechanistic studies to understand how the inhibition is happening,” adds Krogan.
This research will continue under the BioFulcrum Viral and Infectious Disease Research Program at Gladstone and Host Pathogen Mapping Initiative at UCSF.