Binding of SARS-CoV-2 nucleocapsid protein to uninfected epithelial cells induces antibody-mediated complement deposition

SARS-CoV-2 infection triggers strong antibody response toward Nucleocapsid-Protein (NP), suggesting extracellular presence beyond its intra-virion RNA binding. Interestingly, NP was found to decorate infected and proximal uninfected cell-surfaces. Here, we propose a new mechanism through which extracellular NP on uninfected cells contributes to COVID-19 pathogenicity. We show that NP binds to cell-surface sulfated linear-glycosaminoglycans by spatial rearrangement of its RNA-binding sites facilitated by the flexible, positively charged, linker. Coating of uninfected lung-derived cells with purified NP attracted anti-NP-IgG from lung fluids and sera collected from COVID-19 patients. The magnitude of this immune recognition was significantly elevated in moderate compared to mild COVID-19 cases. Importantly, binding of anti-NP-IgG present in sera generated clusters that triggered C3b deposition by the classical complement pathway. Heparin analog enoxaparin outcompeted NP-binding, rescuing cells from anti-NP IgG-mediated complement deposition. Our findings unveil how extracellular NP may exacerbate COVID-19 tissue damage, and suggest leads for preventative therapy. Abstract Figure Highlights IgG from patients’ sera target NP-bound cells resulting in complement activation The flexible linker allows NP to both bind linear sulfated GAGs and wrap around RNA Heparin analogs prevent NP surface binding and alleviate complement activation Cell-ELISA anti-NP IgG levels differ between mild and moderate COVID-19