Video: Said Sannuga, Cellscape.co.uk / ETH Zurich, The Ban Lab
Viruses use the resources of an infected cell to replicate and further infect other cells, and through the same mechanism, the disease spreads to other individuals. The most important stage in the life cycle of a virus is the synthesis of new viral proteins, built on the basis of instructions from the viral RNA genome. The cellular protein factory, the ribosome, begins to rivet viral proteins according to these blueprints.
When the cell is healthy, the ribosome moves along the RNA at a strictly measured pace, reading three RNA nucleotides per step. This three-letter code identifies the corresponding amino acid that attaches to the growing protein. It rarely happens that the ribosome has moved one or two RNA nucleotides forward or backward, departing from this correct three-part pattern. If such a shift in the ribosome (it is called "reading frame shift") does occur, then it leads to errors in reading the genetic code.
In our cells, frame shifting almost never happens; this would render cellular proteins dysfunctional. But the life cycle of some viruses, for example, coronaviruses and HIV, depends precisely on the shift of the reading frame - during such events, the level of viral proteins is regulated. For example, the SARS-CoV-2 virus, the very one that causes COVID-19, is critically dependent on the shift of the reading frame provided by the unusual and confusing folding in the viral RNA.
Therefore, since frame shift is vital for the virus, and almost never occurs in our body, any compound that prevents such a shift could potentially help as an effective antiviral drug. True, it is still not known how viral RNA interacts with the ribosome, provoking a shift in the reading frame, and this would be important to know for drug development.
A detailed picture of the process critical to the replication of the coronavirus obtained
A team of researchers from the Higher Technical School of Zurich and from the Universities of Bern, Lausanne and Cork (Ireland) was for the first time able to identify the interactions between the viral genome and the ribosome that occur during the reading frame shift. The results of their work were published in the journal Science .
With the help of filigree biochemical experiments, the researchers were able to find the ribosome exactly at that site of the SARS-CoV-2 RNA genome, where the reading frame shifted. They managed to study this molecular complex using cryoelectron microscopy.
Based on the results of the work, it was possible to describe this process in unprecedented detail and discover a number of new properties that were not even suspected of. When the reading frame shifts, the ribosomal machine, which is notable for its dynamism, takes on an unnatural configuration, which has helped to obtain one of the sharpest and most accurate images of the mammalian ribosome, visualized during the frame shift, when information is being read from the viral genome. Scientists then traced their structural findings to in vitro and in vivo experiments. and, in particular, how the desired chemical compounds can be targeted at this process. Nenad Ban, Professor of Molecular Biology at the Graduate School of Technology Zurich and co-author of this study, emphasizes that "the results presented here regarding SARS-CoV-2 will also be useful for understanding the mechanisms of reading frameshift in other RNA viruses."
Potential target for antiviral drug development
The dependence of SARS-CoV-2 on such a frame shift event in the ribosome may be useful in the development of antiviral drugs. According to earlier studies, several drugs are known that can inhibit the reading frame shift in coronaviruses; but new research now provides insight into how these compounds affect SARS-CoV-2 levels in infected cells.
In the experiments set up, both compounds suppressed viral replication by two to three orders of magnitude, while not being toxic to the treated cells. At the same time, one of them suppressed viral replication, inhibiting the ribosomal shift of the reading frame, and the action of the second may be based on a different mechanism.
Although these compounds are currently not potent enough to be used therapeutically, this study demonstrates that inhibition of ribosomal frame shift dramatically affects viral replication, which paves the way for the development of better compounds. Due to the fact that all coronaviruses depend on the preservation of such a mechanism for shifting the reading frame, a drug aimed at combating this process may have a therapeutic effect in the fight against other coronaviruses remotely related to SARS-CoV-2. “Our future work will focus on understanding the cellular defense mechanisms that suppress the viral reading of the frame shift, as this may be useful for the development of small compounds with a similar mechanism of action,” says Ban.
The RNA (yellow) of the SARS-CoV-2 virus forms a pseudonodular structure (multicolored, bottom right), which causes a shift when the reading frame moves along the ribosome (brown). Thus, viral RNA controls the levels of viral protein synthesis. More details about the study can be found in the video linked above (Graphics: Said Sannuga, Cellscape.co.uk / ETH Zurich, The Ban Lab)
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