Plasmin‑mediated cleavage between SARS-CoV-2 spike protein and ENaC drives pulmonary edema

The high infectivity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) stems from the insertion of four distinct amino acid residues (RRAR), called polybasic cleavage site in the spike (S) protein, that is strikingly identical with that of epithelial sodium channel (ENaC), both of which can be cleaved by plasmin. Dysregulation of human ENaC, a protein responsible for regulating fluid balance, is directly associated with mortality resulting from pulmonary damage, whereas the involvement of ENaC and regulation by fibrinolytic system during SARS-CoV-2 infection remains elusive. In this study, S plasmid and pseudovirus were applied to mimic SARS-CoV-2 infection, respectively. Pulmonary edema manifestations and dysregulation of ENaC were observed in mice expressing human angiotensin converting enzyme 2, and mechanistic studies revealed both S protein and ENaC could be cleaved by plasmin. Furthermore, our results demonstrated that S protein competitively inhibited the cleavage of ENaC by plasmin, overexpression of ENaC suppressed plasmin-induced cleavage of the S protein, thereby impeding SARS-CoV-2 invasion. Notably, co-enabling the polybasic cleavage site mutated S plasmid/pseudovirus with plasmin reversed the downregulation of ENaC. In conclusion, our data suggest a competitive mechanism of ENaC by SARS-CoV-2 via plasmin was mainly regulated at polybasic cleavage site, indicating a novel mechanism underlying SARS-CoV-2-induced pulmonary edema, and identifying ENaC as a potential therapeutic target for treating SARS-CoV-2-associated respiratory failure.