Researchers at the University of Maryland, Baltimore County (UMBC) have discovered a key step that enteroviruses use to replicate inside human cells. The survey results are nature communicationsexplains how viruses that cause diseases such as polio, encephalitis, myocarditis, and even the common cold control cellular machinery to copy themselves. Scientists say the discovery could ultimately help researchers develop a new generation of antiviral drugs that can target many enteroviruses at once.
The research was led by Deepak Koirala, associate professor of chemistry and biochemistry at UMBC and recent Ph.D. Graduate of Nava Krishna Das. Their research helps answer long-standing questions about how these viruses begin replicating once they enter cells.
“My lab has been working very hard to understand how RNA viruses make proteins inside cells and multiply their genomes to make more virus particles,” Koirala says. The team’s previous work had identified a key cloverleaf-shaped structure within the virus’s RNA. New research shows how that structure recruits proteins needed to build the viral replication machinery.
How enteroviruses replicate within cells
Enteroviruses have very small RNA genomes that need to perform two jobs at once. Viral RNA must both direct the production of viral proteins and serve as a template for making new copies of the virus.
Although most of the viral genome contains instructions for structural proteins, it also encodes several specialized proteins required for replication. One of the most important is a fusion protein called 3CD.
The 3C moiety cleaves long chains of amino acids to produce individual proteins needed by the virus. The 3D part acts as RNA polymerase, an enzyme that copies the viral RNA so the virus can replicate. Human cells do not naturally contain this type of polymerase, so the virus must supply its own version.
“Until now we have determined the structure of only RNA, and other groups have determined 3C and 3D structures, but this time we captured the structure of RNA and protein together, so we can see how they interact,” Koirala explains. “We discovered that it is the 3C domain of 3CD that binds to RNA within the viral genome, which then recruits other components, such as the host protein PCBP2, to assemble the replication complex.”
The researchers also discovered that this molecular complex acts like a switch. Once the 3CD is attached, the virus copies the RNA genome. When the protein is stripped off, the RNA becomes available to make viral proteins in its place.
Scientists unravel long-standing virus mysteries
To examine these interactions in detail, the research team used X-ray crystallography to visualize RNA cloverleaf and 3CD proteins together. They also relied on isothermal titration calorimetry (ITC), which measures the heat released when molecules bind, and biolayer interferometry (BLI), which uses changes in the interference of light to track how long molecules are bound.
This experiment helped settle an ongoing scientific debate. The researchers showed that two complete 3CD molecules, each carrying its own RNA polymerase, bind side-by-side on the viral RNA. Previous studies proposed that the proteins form a single fusion pair.
Scientists still don’t fully understand why two copies are needed, but new research provides a clearer picture of how the replication process begins.
Potential for broad-spectrum antivirals
One of the most promising findings was how similar the mechanisms were in all seven enteroviruses examined in the study. These viruses shared nearly identical RNA clover structure and binding behavior.
This level of similarity suggests that RNA structure is critical for virus survival. Critical mutations are likely to disrupt replication, potentially making the structure a stable drug target across many enteroviruses.
Researchers say this raises the possibility of developing broad-spectrum antivirals that act on entire families of viruses rather than single pathogens.
Scientists have already developed drugs to interfere with 3C and 3D proteins, but new discoveries reveal another possible strategy.
Drugs that block the activity of 3C and 3D are already in development, but “now we have another layer to test,” Koirala says. “What if we targeted the RNA, or the RNA-protein interface, to disrupt the interaction? That’s another opportunity. Now that we have high-resolution structures, we can precisely design drug molecules to target them.”
Koirala said the study highlights how surprisingly sophisticated the virus is despite its small genome.
“Viruses are very smart. Their entire genome is equivalent to about one human mRNA sequence, but they are very effective,” Koirala says. His latest research demonstrates “why we need to study this basic science so that it can be applied to the development of drugs that target the pathogens that cause so many harmful diseases.”
