Factor Xa cleaves SARS-CoV-2 spike protein to block viral entry and infection.

Serine proteases (SP), including furin, trypsin, and TMPRSS2 cleave the SARS-CoV-2 spike (S) protein, enabling the virus to enter cells. Here, we show that factor (F) Xa, an SP involved in blood coagulation, is upregulated in COVID-19 patients. In contrast to other SPs, FXa exerts antiviral activity. Mechanistically, FXa cleaves S protein, preventing its binding to ACE2, and thus blocking viral entry and infection. However, FXa is less effective against variants carrying the D614G mutation common in all pandemic variants. The anticoagulant rivaroxaban, a direct FXa inhibitor, inhibits FXa-mediated S protein cleavage and facilitates viral entry, whereas the indirect FXa inhibitor fondaparinux does not. In the lethal SARS-CoV-2 K18-hACE2 model, FXa prolongs survival yet its combination with rivaroxaban but not fondaparinux abrogates that protection. These results identify both a previously unknown function for FXa and an associated antiviral host defense mechanism against SARS-CoV-2 and suggest caution in considering direct FXa inhibitors for preventing or treating thrombotic complications in COVID-19 patients.

Potent NKT cell ligands overcome SARS-CoV-2 immune evasion to mitigate viral pathogenesis in mouse models.

One of the major pathogenesis mechanisms of SARS-CoV-2 is its potent suppression of innate immunity, including blocking the production of type I interferons. However, it is unknown whether and how the virus interacts with different innate-like T cells, including NKT, MAIT and γδ T cells. Here we reported that upon SARS-CoV-2 infection, invariant NKT (iNKT) cells rapidly trafficked to infected lung tissues from the periphery. We discovered that the envelope (E) protein of SARS-CoV-2 efficiently down-regulated the cell surface expression of the antigen-presenting molecule, CD1d, to suppress the function of iNKT cells. E protein is a small membrane protein and a viroporin that plays important roles in virion packaging and envelopment during viral morphogenesis. We showed that the transmembrane domain of E protein was responsible for suppressing CD1d expression by specifically reducing the level of mature, post-ER forms of CD1d, suggesting that it suppressed the trafficking of CD1d proteins and led to their degradation. Point mutations demonstrated that the putative ion channel function was required for suppression of CD1d expression and inhibition of the ion channel function using small chemicals rescued the CD1d expression. Importantly, we discovered that among seven human coronaviruses, only E proteins from highly pathogenic coronaviruses including SARS-CoV-2, SARS-CoV and MERS suppressed CD1d expression, whereas the E proteins of human common cold coronaviruses, HCoV-OC43, HCoV-229E, HCoV-NL63 and HCoV-HKU1, did not. These results suggested that E protein-mediated evasion of NKT cell function was likely an important pathogenesis factor, enhancing the virulence of these highly pathogenic coronaviruses. Remarkably, activation of iNKT cells with their glycolipid ligands, both prophylactically and therapeutically, overcame the putative viral immune evasion, significantly mitigated viral pathogenesis and improved host survival in mice. Our results suggested a novel NKT cell-based anti-SARS-CoV-2 therapeutic approach.

SARS-CoV-2 Nsp5 Demonstrates Two Distinct Mechanisms Targeting RIG-I and MAVS To Evade the Innate Immune Response.

Newly emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic with astonishing mortality and morbidity. The high replication and transmission of SARS-CoV-2 are remarkably distinct from those of previous closely related coronaviruses, and the underlying molecular mechanisms remain unclear. The innate immune defense is a physical barrier that restricts viral replication. We report here that the SARS-CoV-2 Nsp5 main protease targets RIG-I and mitochondrial antiviral signaling (MAVS) protein via two distinct mechanisms for inhibition. Specifically, Nsp5 cleaves off the 10 most-N-terminal amino acids from RIG-I and deprives it of the ability to activate MAVS, whereas Nsp5 promotes the ubiquitination and proteosome-mediated degradation of MAVS. As such, Nsp5 potently inhibits interferon (IFN) induction by double-stranded RNA (dsRNA) in an enzyme-dependent manner. A synthetic small-molecule inhibitor blunts the Nsp5-mediated destruction of cellular RIG-I and MAVS and processing of SARS-CoV-2 nonstructural proteins, thus restoring the innate immune response and impeding SARS-CoV-2 replication. This work offers new insight into the immune evasion strategy of SARS-CoV-2 and provides a potential antiviral agent to treat CoV disease 2019 (COVID-19) patients. IMPORTANCE The ongoing COVID-19 pandemic is caused by SARS-CoV-2, which is rapidly evolving with better transmissibility. Understanding the molecular basis of the SARS-CoV-2 interaction with host cells is of paramount significance, and development of antiviral agents provides new avenues to prevent and treat COVID-19 diseases. This study describes a molecular characterization of innate immune evasion mediated by the SARS-CoV-2 Nsp5 main protease and subsequent development of a small-molecule inhibitor.

Mathematical modeling of interaction between innate and adaptive immune responses in COVID-19 and implications for viral pathogenesis.

We have applied mathematical modeling to investigate the infections of the ongoing coronavirus disease-2019 (COVID-19) pandemic caused by SARS-CoV-2 virus. We first validated our model using the well-studied influenza viruses and then compared the pathogenesis processes between the two viruses. The interaction between host innate and adaptive immune responses was found to be a potential cause for the higher severity and mortality in COVID-19 patients. Specifically, the timing mismatch between the two immune responses has a major impact on disease progression. The adaptive immune response of the COVID-19 patients is more likely to come before the peak of viral load, while the opposite is true for influenza patients. This difference in timing causes delayed depletion of vulnerable epithelial cells in the lungs in COVID-19 patients while enhancing viral clearance in influenza patients. Stronger adaptive immunity in COVID-19 patients can potentially lead to longer recovery time and more severe secondary complications. Based on our analysis, delaying the onset of adaptive immune responses during the early phase of infections may be a potential treatment option for high-risk COVID-19 patients. Suppressing the adaptive immune response temporarily and avoiding its interference with the innate immune response may allow the innate immunity to more efficiently clear the virus.