Introduction
COVID-19 has remained a major health concern since the World Health Organization (WHO) declared it a pandemic on 11 March 2020 (1). As of 16 May 2021, severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2) has infected more than 161 million people worldwide, with over 3 million deaths (2). This has imposed a huge burden on health care systems worldwide and provoked a serious economic crisis following inevitable social lockdown (3). Thus, there is a desperate need for developing sensitive and robust diagnostic tests and potentially effective vaccines for SARS-CoV-2 to alleviate the growing socioeconomic impacts. To this goal, obtaining illuminating insights into the immune response, particularly the antibody response, to SARS-CoV2 is obviously crucial. From the experience with SARS-CoV virus, it is postulated that neutralizing antibodies elicited against SARS-CoV-2 in convalescent individuals may be important correlates of protection (aroused either by natural infection or vaccination) (4, 5), and detection of virus-specific antibodies may provide a robust diagnostic tool for epidemiological purposes. Moreover, early seroconversion and high antibody titers could be used as a prognostic marker for disease severity and ARDS development in patients with SARS-CoV-2 (5). However, despite extensive investigations, precise definition of the immune responses, especially targets of adaptive immune responses to SARS-CoV-2 infection is still elusive.
Like other coronaviruses infecting humans, SARS-CoV-2 is an enveloped positive-sense RNA virus, composed of four structural proteins known as spike (S), envelope (E), membrane (M), and nucleocapsid (NP) proteins (6). Apart from NP which is associated to the viral RNA genome, the other three, S, E, and M proteins organize the viral envelope and are directly accessible to the host immune system (6). The S protein is a large type I transmembrane glycoprotein that is responsible for receptor binding and membrane fusion. With a total length of 1273 amino acids, S protein has two functional domains, S1 and S2, located in the N- and C-terminal, respectively (7, 8). S1 subunit itself consists of an N-terminal domain (14–305 residues) and a receptor-binding domain (RBD, 319–541 residues) which binds to the host receptor angiotensin converting enzyme 2 (ACE2), while the S2 subunit which plays an important role in SARS-CoV-2 fusion with the target cells, harbors the fusion peptide (FP) (788–806 residues), heptapeptide repeat sequence 1 (HR1) (912–984 residues), HR2 (1163–1213 residues), transmembrane (TM) domain (1213–1237 residues), and cytoplasmic domain (1237–1273 residues) (8, 9). It has become evident that SARS-CoV-2 like other human-infecting coronaviruses (e.g. SARS-CoV, MERS-CoV) can elicit IgM and IgG antibodies directed against the S and NP protein, and the RBD of S protein, after initial infection (10). This antibody response is initially used in serological assays for diagnostic purposes (11). However, due to the presence of several cross-reactive epitopes in NP and S with SARS-CoV and MERS-CoV (12), the detected antibody response may not represent only anti-SARS-CoV-2 antibodies and could be a reflection of previous exposures to other human coronaviruses (13). Moreover, these serology-based assays do not offer any clue about antibody functionality, including neutralization potential which is a prerequisite in vaccine development. On the other hand, antibody-dependent enhancement (ADE) through low quality, low quantity, and non-neutralizing antibodies is a concern with human coronaviruses (14), further necessitates epitope dissection in order to resolve virus-specific and favorable antibodies from unfavorable cross-reactive antibodies.
Since SARS-CoV-2 virus largely depends on the spike glycoprotein for binding to ACE2 receptor and cell entry (6), several studies have focused on detecting antibodies against S protein and its RBD to assess their neutralizing effect. However, antibodies against nucleoprotein appear earlier in the course of seroconversion (15) and could be more indicative for diagnostic applications. In the current study, we present several immunodominant epitopes on the S and NP proteins by Pepscan analysis on the sera from Iranian SARS-CoV-2 patients. This information may be useful for designing COVID-19 diagnostics and vaccines.
Materials and Methods