Molecular Interactions Between SARS-CoV-2 Viral and Human Cellular Factors: Implications for Viral Infection, Replication, and Disease

Abstract

The COVID-19 pandemic was this century's most significant global public health crisis. The causative agent was SARS-CoV-2, an enveloped, single-stranded positive-sense virus. As an obligate intracellular pathogen, SARS-CoV-2 hijacks host cellular machinery to complete its life cycle while evading or counteracting the host immune response. Understanding the molecular interactions between host factors and viral proteins is crucial for comprehending COVID-19 pathogenesis and developing new antiviral strategies. Objective I: Identification and Characterization of Cellular Targets for Host-Directed Therapy Against SARS-CoV-2 Numerous research groups have utilized high-throughput methodologies to profile host responses during SARS-CoV-2 infection, generating extensive data on virus-host interactions. Our goal was to identify consistently upregulated host factors at infection sites and understand their roles in viral infection and disease progression. We conducted a meta-analysis of published transcriptome and proteome profiles from nasal swab and bronchoalveolar lavage fluid (BALF) samples of COVID-19 patients, identifying Thioredoxin as a consistently upregulated host factor that is involved in cellular Redox regulation. The active and inactive thioredoxin balance is maintained by thioredoxin reductase, which can be inhibited by the FDA-approved drug Auranofin. We tested Auranofin’s antiviral efficacy in vitro and in vivo using preclinical Syrian golden hamsters, establishing that the drug's protective mechanism includes targeting viral entry, protease activity, and suppressing the proinflammatory cytokine IL-6 production. Objective II: Characterization of Cellular Antiviral Responses Against SARS-CoV-2 and Viral Evasion Strategies Cellular intrinsic innate immunity is critical in restricting viral infection and replication. It is driven by Interferon induction and signaling, leading to the expression of antiviral host factors that create an antiviral state at the site of infection. We aimed to identify SARS-CoV-2 proteins that antagonize the interferon (IFN) induction and signaling pathways and elucidate their molecular mechanisms. IFN reporter-based screening revealed that ORF6, among other viral proteins, is a potent inhibitor of IFN induction and signaling. This activity was mapped to the C-terminal cytoplasmic tail, specifically amino acid residues 52–61. ORF6 was found to directly interact with RIG-I, reducing K63-linked ubiquitination of RIG-I and thereby inhibiting downstream type I IFN induction and signaling. This involves ORF6-mediated degradation of TRIM25, a process observed during SARS-CoV-2 infection. Overall, our findings provide new cellular targets for host-directed therapy for COVID-19 and provide novel insights into innate immune evasion by SARS-CoV-2.