Lab experiments add further evidence to bradykinin storm theory of viral pathogenesis of COVID-19

A new article published in Nature Communication adds further evidence to bradykinin storm theory of viral pathogenesis of COVID-19 -; a theory that was put forward two years ago by a team of researchers at the Department of Energy’s Oak Ridge National Laboratory.

At the height of the pandemic, ORNL systems biologist Dan Jacobson and his team used ORNL’s Summit supercomputer to analyze gene expression data from lung cells of COVID-19 patients. Their research suggested that genes related to some of the body systems that are responsible for controlling blood pressure, fluid balance and inflammation appear to be excessively dysregulated or altered in the lung cells of people infected with the virus. In an article published in eLife, the team predicted that the overproduction of bradykinin -; the compound that dilates blood vessels and makes them permeable -; could be the source of COVID-19 symptoms such as excessive fluid accumulation in the lungs, fatigue, nausea and decreased cognitive function.

This theory has been supported by a new study led by Jacobson and his colleagues from ORNL’s Biosciences, Computational Sciences and Engineering, and Neutron Scattering divisions in collaboration with Soichi Wakatsuki, professor of photon science at the University’s SLAC National Accelerator Laboratory. from Stanford. Wakatsuki’s team was able to experimentally prove that the virus’ main protease, 3CLpro, binds to the essential modulator NF-κB, or NEMO. The subsequent cleavage of NEMO means that it deregulates NF-κB, which is a protein complex that helps regulate the immune system’s response to infection -; and its dysregulation may contribute to a bradykinin storm, just as the ORNL team’s model of pathogenesis predicted.

“It’s the culmination of a lot of work from a lot of different angles,” Jacobson said. “We’re a computational systems biology group, so our previous work was really based on large-scale data analysis. It takes all that computational work in the wet lab to generate new datasets to confirm activity. enzymatic and structural interactions. It’s incredibly exciting to see all of these lines of evidence coming together and then being validated – that everything our previous work predicted to be the case is actually true.”

At SLAC, Wakatsuki’s team was able to use viral3CLpro proteins (produced by ORNL senior scientist Andrey Kovalevsky) and peptides to represent cleavage sites in NEMO. The team then used X-ray crystallography to show the structural interaction between the two. Additionally, an ORNL team led by former ORNL researcher Stephanie Galanie was able to show, biochemically, that 3CLpro can cleave NEMO at physiologically relevant concentrations.

We now have evidence at the atomistic level and biochemistry supporting the hypothesis that it binds and cleaves exactly as we expected.”

Dan Jacobson, ORNL Systems Biologist

This inter-laboratory collaboration at ORNL and SLAC originated through the National Virtual Biotechnology Laboratory, or NVBL, a DOE program funded by the Coronavirus Aid, Relief and Economic Security Act in 2020, which encouraged national laboratories in the fight against COVID-19. Wakatsuki and Jacobson met after Jacobson gave a presentation at one of NVBL’s virtual sessions and asked collaborators to help him prove his bradykinin storm theory through biology experiments. structural.

“We went to get people to do this next step with us, and Soichi spoke up at one of the meetings and said, ‘Yeah, let’s go. And here we are now with a beautiful high-impact article. I think that’s a real benefit of the collaborative approach the NVBL has had the national labs working on, and I’d love to see more of that,” Jacobson said.

As part of this effort, ORNL computational systems biologist Erica Prates, then a postdoctoral researcher and now an early-career staff member in the Biosciences Division, coordinated a team including Omar Demerdash, Julie Mitchell, and Stephan Irle of the ORNL. They conducted extensive molecular dynamics work on Summit using both quantum mechanics and machine learning methods to examine the binding affinity of NEMO and 3CLpro in humans and other species and to examine the structural models derived from sequences of other coronaviruses.

“Erica plays an important role in what we call structural systems biology to bridge computational efforts in the fields of systems biology and structural biology,” Jacobson said.

This team’s research will provide a better understanding of the effects of different viruses, including zoonotic diseases, which are human diseases of animal origin, in different host species. This knowledge will be essential in the effort to predict or even prevent the next pandemic.

“Our COVID work continues, but much of our focus has shifted to pandemic prevention,” Jacobson said. “We have new funding secured in conjunction with a number of other research institutions that are really focusing on dynamic prevention and trying to understand the rules of zoonosis and the effects of, for example, climate change and how it drives new zoonotic overflow events.

Jacobson and his colleagues are partnering with Johns Hopkins University, Cornell University and others to conduct a wide range of studies and field trials to analyze the interactions between viral proteins and host proteins, creating the datasets needed for computer models that will make predictions of viruses across a range of species.

“Why do viruses happily live in some species nonpathogenically, but become pathogenic when zoonosis spread occurs? How do they hop between different host species and are nonpathogenic until they hit humans? said Jacobson. “The rules behind zoonosis are very poorly understood, and we have some really exciting work going on where we’re building predictive models to understand the variables in the environment that can lead to these spillover events.”

The teams’ research was also partially funded by ORNL’s laboratory-led research and development program, which supported conceptual work on NEMO cleavage in animal models for COVID-19 pathology. This work utilized DOE Office of Science user facilities, including the Oak Ridge Leadership Computing Facility, Spallation Neutron Source and High Flux Isotope Reactor, all at ORNL, and the Stanford Synchrotron Radiation Lightsource at SLAC.

Funding for the conceptualization of human pathogenesis was provided by a grant from the National Institutes of Health.


Journal reference:

Hamedi, MA, et al. (2022) Structural and functional characterization of NEMO cleavage by SARS-CoV-2 3CLpro. Nature Communication.

Sharon D. Cole