Rotavirus is one of the main pathogens that cause diarrhea in infants and young children. It mainly infects intestinal epithelial cells, causing cell damage and causing diarrhea. It causes 130,000 infants and young children under the age of 5 to die every year. It is the single main cause of diarrhea in infants and young children. Almost every child in the world about five years old has been infected with rotavirus at least once.
Viroplasms have been difficult to study because they usually form very quickly, but an accidental observation allowed researchers at Baylor College of Medicine to discover new insights into the formation of viral plastids.
The researchers created a mutant rotavirus whose replication speed was unexpectedly much slower than the original virus, which allowed them to observe the first step of virus assembly.
The discovery, published in the Journal of Virology, opens up new possibilities for treating and preventing this viral disease and understanding how similar factories work with other viruses.
“The formation of viral virulence is essential for the successful infection of rotavirus. They form rapidly in infected cells, are composed of viruses and cellular proteins, and interact with lipid droplets, but how are these components combined? The details are still unclear,” said first author Jeanette M. Criglar, a former postdoctoral intern and a scientist in the laboratory of Dr. Mary Estes in the Department of Molecular Virology and Microbiology at Baylor.
Lipid droplets are complex and active multifunctional organelles. Their basic functions are mainly involved in lipid synthesis and storage. Lipid-related proteins participate in various lipid-related biological processes, including cell signal transduction, regulation of lipid metabolism, Membrane transport, synthesis and secretion of inflammation-related proteins, etc.
Many pathogens, including viruses, intracellular bacteria and protists, target lipid droplets. Pathogenic microorganisms can induce the increase of intracellular lipid droplets, and the increased lipid droplets can provide a platform for the proliferation of pathogenic microorganisms.
In order to gain a new understanding of the formation of viral plastids, Criglar and her colleagues studied NSP2, a viral protein necessary for viral replication. Without it, neither virus virulence nor new viruses will form.
Like all proteins, NSP2 is composed of amino acids, which are strung together like beads on a necklace. ‘Bead’ 313 is serine. Importantly, serine 313 is phosphorylated-it has a phosphate group attached to it.
Protein phosphorylation is a mechanism by which cells regulate protein activity. It works like a switch to activate or deactivate proteins.
Here, the researchers evaluated the role of NSP2 phosphorylated serine 313 in the formation of virions.
Criglar and her colleagues used the recently developed reverse genetics system by converting serine to aspartic acid to produce a rotavirus with the NSP2 protein. This virus has a 313 amino acid mutation called Phosphorus mutation.
The name Phosphoprotein indicates that the mutant protein mimics the phosphorylated protein in the original rotavirus.
Reverse genetics starts with a protein, works in reverse, produces mutant genes, and then makes them part of a virus to study the effect of the protein on the behavior of the virus.
Criglar said: “In laboratory experiments, our phosphoric mutant protein crystallized faster than the original protein, within a few hours instead of a few days.”
“But surprisingly, compared to non-mutated rotaviruses, the phosphoroviruses produce virulence and replicate very slowly.”
“This is not what we expected. We think rotavirus with mutant proteins will also replicate faster.”
“We used the delay in the formation of virions to observe early events that are difficult to study.”
The researchers found that the first step in the formation of virions is the binding of NSP2 to lipid droplets, which indicates that NSP2, which is phosphorylated only at position 313, can interact with lipid droplets and not with other components of the virulence.
Lipid droplets are an important part of viral toxicants. It is well known that rotavirus tricks infected cells to produce lipid droplets, but how it is done is not clear.
New findings suggest that rotavirus may use phosphorylated NSP2 to trigger the formation of lipid droplets.
Criglar said: “It is very exciting to see that changing only one amino acid in the NSP2 protein affects the replication of the entire virus.”
“This change in phosphorus-like changes the dynamics of virus replication without killing the virus. We can use this mutant rotavirus to continue to study the sequence of events that lead to the formation of virions, including a long-standing cell biology biology. Questions about how lipid droplets are formed.”