NSP1 was found to act as an antigenic factor for SARS-CoV-2 infections

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has had a major impact on global health, social and economic well-being for over two years, following the unexpected appearance in December 2019. ..

This beta coronavirus is transcribed in the host cell and cleaved by viral proteases to produce multiple polyproteins that produce structural and nonstructural proteins.

A new study is investigating the role of nonstructural protein 1 (NSP1) in SARS-CoV-2 infection and etiology.

Preface

Viruses are intracellular parasites that hijack host cell mechanisms to convert their genetic code into proteins, driving further steps in the replication and transmission of infections. Normally, they shut down the translation of the host protein, also known as “host shut off”. This helps to take over cells by rerouting the protein translation mechanism from cellular messenger ribonucleic acid (mRNA) to viral mRNA, while at the same time avoiding the immune response of host cells targeting the virus. increase.

The virus uses a number of strategies to achieve host blockade, including direct blockade of translation, disruption of mRNA transcription and transport from the nucleus to the cytoplasm, and indirect methods such as inducing mRNA degradation. To do. especially, coronavirus (CoVs) have been studied on this aspect.

The authors of this study have already shown that all three of the above mechanisms function in SARS-CoV-2 infection. Interestingly, NSP1 is the most well-understood host blocker and is also important in the etiology of betaCoV.

The early SARS-CoV pathogen, NSP1, previously blocked host protein translation through a number of mechanisms, including blocking mRNA invasion channels, suppressing cellular mRNA exports, and facilitating cellular mRNA degradation. Shown in the experiment. This latter does not affect viral proteins due to the 5’leader sequence, but the interaction between each of these processes can indirectly affect each of the other processes.

In the current paper, it is published in the journal Cell reportNSP1 mutants were overexpressed to understand how this protein affects the expression of cellular genes.

Survey results

Researchers have found that NSP1 acts on host cells by increasing the rate of cellular mRNA degradation and at the same time blocking the translation of host proteins. Wild-type NSP1 causes endonuclease cleavage of cellular mRNA, making them more prone to degradation. Both of these processes appear to be dependent on NSP1-ribosome binding, as in SARS-CoV.

Furthermore, the mutant NSP1 retained the ability to block translation of the host RNA, but lost the ability to accelerate RNA degradation, indicating that these two processes clearly occur. NSP1 also seems to inhibit its own expression.

NSP1 also interacts with the NXF1 protein via the N-terminal region, interfering with the export of nuclear mRNA. The location of this interaction seems to indicate that it is independent of the aforementioned ribosome-NSP1 binding. In other words, NSP1 is “You can target three different steps of the mRNA biosynthetic pathway.. “

Another finding was that NSP1 induced degradation of cytosolic RNA in infected cells, as indicated by a significant reduction in the half-life of cytosolic transcripts that bind to ribosomes compared to nuclear transcripts. .. This has nothing to do with curbing nuclear RNA exports.

Induction of mutant NSP1 with a deletion of amino acids 155 to 165 did not reduce protein expression, but the mutant appeared to show stronger binding to nuclear RNA export proteins. However, if wild and mutant SARS-CoV-2 viruses are allowed to infect cells with an interferon (IFN) antiviral response during operation, the mutant virus will be other over a long period of observation. It showed a reduction in proliferation compared to the virus.

Since the IFN-stimulated gene (ISG) was more strongly induced in mutant-infected cells compared to wild-type infected cells, reduced expression of the viral gene is likely the result of NSP1 interference with the interferon response. Thus, inhibition of proliferation of mutant-infected cells was caused by increased intensity of antiviral type I IFN response, as evidenced by higher IFNꞵ protein levels compared to wild-type virus-infected cells.

Again, ruxolitinib, a selective JAK-STAT inhibitor that suppresses the IFN pathway, rescued the mutant virus from its replication blockade.

These results indicate that enhanced IFN response in cells infected with CoV2-mut is responsible for their proliferative disorders... “

This study also showed that NSP1 is the main driver of host blockade after SARS-CoV-2 infection through its involvement with the ribosome. Viral genes also showed low translational efficiency (TE) comparable to the lower end of cellular genes. This was probably due to the corridor of large amounts of viral RNA in the replication compartment where the ribosomes could not access them.

The viral gene TE is further affected by the presence of mutations in NSP1 and indicates that wild-type NSP1 increases viral protein synthesis through its ribosome involvement. Nevertheless, protein translation levels are reduced by a factor of four in wild-type and mutant-infected cells. This strongly suggests that mutant NSP1 results in equivalent or better levels of translation compared to wild-type proteins.

Mutant NSP1 allows the production of viral RNA at constant levels, but does not block the host RNA. However, cytosol transcripts were found to be more strongly reduced in wild-type viruses compared to NSP1 mutants. The half-lives of these cytosol transcripts were also low. This indicates NSP1-dependent degradation of cellular transcripts mediated by NSP1 ribosome binding.

This is supported by increased intron readings due to wild-type SARS-CoV-2 infection compared to mutant infection due to the high degradation rate of mature cytoplasmic sol RNA after infection.

In vivo Hamster studies also showed that, unlike wild-type viruses, NSP1 mutants did not cause significant weight loss after infection, demonstrating an important pathogenic role for proteins. Within a week of infection, weight was significantly reduced, but those infected with the wild-type virus had higher lung and nasal mucosal viral titers compared to mutants. As expected, ISG was upregulated in the latter lung and cytosol RNA was expressed at low levels.

Implications

This striking study reveals the central role of NSP1 in blocking protein synthesis after SARS-CoV-2 infection and reveals three major mechanisms of action. Of these, inhibition of RNA degradation and translation appears to be the result of interaction with the ribosome, whereas inhibition of nuclear RNA export is independent of ribosome binding.

Viral genes avoid NSP1-dependent inhibition of translation of both viral and cellular mRNA. However, because the latter is degraded by NSP1, the available ribosomes now translate only viral RNA. Alternatively, or in addition, viral mRNA may resist suppression by NSP1 and release ribosome entry channels due to conformational changes induced by this protein after binding.

Host blockade can promote viral growth, increase viral protein translation, and suppress the antiviral response of host cells. The results of this study indicate that the latter is more important of the two effects, as the antiviral type I IFN response and increased ISG response in infected cells and hamsters led to a reduction in the growth of mutant viruses. Is shown. No host shutoff.

Overall, our data demonstrate that nsp1 is the major antigenic escape factor for SARS-CoV-2... “