Abstract
COVID-19 is currently a highly pressing health threat and therapeutic strategies to mitigate the infection impact are urgently needed. Characterization of the SARS-CoV-2 interactome in infected cells may represent a powerful tool to identify cellular proteins hijacked by viruses for their life cycle and develop host-oriented antiviral therapeutics. Here we report the proteomic characterization of host proteins interacting with SARS-CoV-2 Nucleoprotein in infected Vero E6 cells. We identified 24 high-confidence proteins mainly playing a role in RNA metabolism and translation, including RNA helicases and scaffold proteins involved in the formation of stress granules, cytoplasmic aggregates of messenger ribonucleoproteins that accumulate as a result of stress-induced translation arrest. Analysis of stress granules upon SARS-CoV-2 infection showed that these structures are not induced in infected cells, neither eIF2α phosphorylation, an upstream event leading to stress-induced translation inhibition. Notably, we found that G3BP1, a stress granule component that associates with the Nucleoprotein, is required for efficient SARS-CoV-2 replication. Moreover, we showed that the Nucleoprotein-interacting RNA helicase DDX3X colocalizes with viral RNA foci and its inhibition by small molecules or small interfering RNAs significantly reduces viral replication. Altogether, these results indicate that SARS-CoV-2 subverts the stress granule machinery and exploits G3BP1 and DDX3X for its replication cycle, offering groundwork for future development of host-directed therapies.
Keywords: Antiviral drugs; COVID-19; DEAD RNA helicase; G3BP1; Proteomics; Stress granules.
【저자키워드】 COVID-19, proteomics, antiviral drugs, DEAD RNA helicase, G3BP1, Stress granules., 【초록키워드】 viruses, Structure, SARS-CoV-2, proteomics, Stress, translation, SARS-COV-2 infection, antiviral drugs, Infection, metabolism, RNA, Replication, Helicase, Protein, Health, Viral, viral replication, Phosphorylation, Therapeutic strategies, VERO E6 cells, Small molecules, antiviral therapeutics, small molecule, Viral RNA, nucleoprotein, stress granule, stress granules, SARS-CoV-2 replication, therapeutic strategy, proteomic, G3BP1, cellular, host proteins, DDX3X, ribonucleoprotein, Vero E6, life cycle, characterization, scaffold proteins, small interfering RNAs, infected cells, host protein, Host-directed therapies, powerful tool, eIF2α, SARS-CoV-2 nucleoprotein, aggregate, infected cell, upstream, foci, infected Vero E6 cells, mitigate, identify, develop, virus, significantly, involved, required, cytoplasmic, reduce, accumulate, granules, the SARS-CoV-2, 【제목키워드】 Proteomic analysis, RNA, Helicase, Host, identify,