SARS CoV-2 Nucleocapsid Protein-mediated Enhancement of Spike pseudotyped Lentivirus Infectivity and Loss of Sensitivity to Neutralisation

Abstract

BACKGROUND: The causative agent of the ongoing COVID-19 pandemic, SARS CoV-2, is a highly pathogenic virus requiring specialized biocontainment facilities. However, pseudotyping-based approaches using the spike glycoprotein have allowed for their study in BSL-2 level laboratories, enabling the rapid screening and identification of neutralizing antibodies, entry inhibitors, host factors, and therapeutic agents. However, this minimalist approach fails to capture the possible contributions and roles of other SARS CoV-2 genes in the entry process. OBJECTIVES: To determine the relative effects of structural and non-structural genes of the SARS CoV-2 on the infectivity of spike-pseudotyped particles, using a lentiviral vector system. METHODS AND RESULTS: An unbiased co-transfection screen of twenty-four SARS CoV-2 genes revealed the nucleocapsid (N) protein as a prime promoter of spike-pseudotyped lentivirus infectivity, as assayed by transduction of an ACE2+ cell line. The spike protein was also observed to be enriched in virions when augmented by the presence of the N gene during virus production. Further, N-enhanced spike-pseudoviruses exhibited a lowered sensitivity to neutralisation by an IgG-Fc fused ACE2 microbody. These results highlight the broad importance of incorporating specific accessory genes during spike-pseudovirus preparation, which may help better recapitulate a physiologically relevant in vitromodel for SARS CoV-2 infectivity.