The Nucleocapsid Protein of SARS–CoV-2: a Target for Vaccine Development

Abstract

KEYWORDS coronavirus disease 2019 (COVID-19), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), nucleocapsid protein, vaccine D uring the current coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), there has been an unprecedented level of global collaboration that has led to a rapid characterization of SARS-CoV-2 (1). Its sequence shares 79.6% identity to SARS-CoV (1, 2), the infectious virus that caused an epidemic in 2003 (2, 3). SARS-CoV-2 has a single-stranded, plus-sense, RNA genome of approximately 30 kb, which includes five major open reading frames encoding nonstructural replicase polyproteins and structural proteins (1), namely, spike (S) (4-6), envelope (E), membrane (M), and nucleocapsid (N) (7), and they are in the same order and of approximately the same sizes as those in SARS-CoV. The SARS-CoV-2 S protein is being used as the leading target antigen in vaccine development (8, 9). However, the complex molecular details of viral entry may lead to complications with the vaccine response, similar to those seen with HIV type 1 (HIV-1) Env protein vaccine efforts (10). The SARS-CoV-2 S gene has 76% amino acid similarity to the SARS-CoV S gene (11), and nonsynonymous mutations developed in the S protein as the SARS-CoV epidemic progressed (12, 13). In contrast, the N gene is more conserved and stable, with 90% amino acid homology and fewer mutations over time (2, 3, 11, 14-16). N proteins of many coronaviruses are highly immunogenic and are expressed abundantly during infection (17). High levels of IgG antibodies against N have been detected in sera from SARS patients (18), and the N protein is a representative antigen for the T-cell response in a vaccine setting, inducing SARS-specific T-cell proliferation and cytotoxic activity (19, 20). We have already shown that the middle or C-terminal region of the SARS-CoV N protein is important for eliciting antibodies against SARS-CoV during the immune response (21-23). New reports have additionally shown that the crystal structure of the SARS-CoV-2 nucleocapsid protein is similar to those of previously described coronavirus N proteins, but their surface electrostatic potential characteristics are distinct (7). Sheikh et al. studied the factors influencing N gene variations among 13 coronaviruses and how these affect virus-host relationships, reporting a high AT% and low GC% in the nucleotide contents of SARS coronavirus (24). In this issue, Cong et al. (17) used a mouse hepatitis virus (MHV) model to show that the viral nucleocapsid (N) protein contributes to forming helical ribonucleoproteins during the packaging of the RNA genome, regulating viral RNA synthesis during replication and transcription and modulating metabolism in infected subjects. This study complements others that have shown N to have multiple functions (25). It is becoming more evident just how critical this protein is for multiple steps of the viral life cycle. These reports offer important and timely insights relevant ...

Head-to-head evaluation on diagnostic accuracies of six SARS-CoV-2 serological assays

“…11 The N protein, on the other hand, has been described to be highly immunogenic and expressed in large quantities during acute infection of SARS-CoV-2. 12 Western blot studies also indicate that the N protein is a potent viral antigen for SARS-CoV-2 on the basis of IgG, IgA and IgM antibodies directed against N protein in COVID-19 positive sera. 13 To our knowledge, our study is the first which investigated and compared immunoassays with different design configurations against the S1 subunit, RBD protein, and N antigen, including that of a surrogate viral neutralising antibody (NAb) assay.…”

Global BioID-based SARS-CoV-2 proteins proximal interactome unveils novel ties between viral polypeptides and host factors involved in multiple COVID19-associated mechanisms

“…Together, the ORF8 BioID interactome did not provide striking clues to narrow down its putative functions in cells. expressed during infection 155,156 . N binds viral RNA through both its Nand C-terminal domains, connected by a disordered SR-rich linkage region 157,158 .…”

SARS-CoV-2: Comparative analysis of different RNA extraction methods

“…However, several labs reported cases in which the N gene was the only gene detected at high Ct (>36) which was, instead, undetectable by re-testing some subjects the day after (unpublished observations). Although conserved among coronaviruses, it has been shown that the N gene exhibited a great variability ( Wang et al, 2020 ; Ceraolo and Giorgi, 2020 ; Dutta et al, 2020 ). Besides the variability of replicate specimens and of the run’s volume reactions, it can be hypothesized that the uncertainty on N positive results could be linked to a transient stage of virus-host contact and its transitory detection to its high abundancy.…”