Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the COVID-19 global pandemic, utilizes the host receptor angiotensin-converting enzyme 2 (ACE2) for viral entry. However, other host factors might also play important roles in SARS-CoV-2 infection, providing new directions for antiviral treatments. GRP78 is a stress-inducible chaperone important for entry and infectivity for many viruses. Recent molecular docking analyses revealed putative interaction between GRP78 and the receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein (SARS-2-S). Here we report that GRP78 can form a complex with SARS-2-S and ACE2 on the surface and at the perinuclear region typical of the endoplasmic reticulum in VeroE6-ACE2 cells and that the substrate-binding domain of GRP78 is critical for this interaction. In vitro binding studies further confirmed that GRP78 can directly bind to the RBD of SARS-2-S and ACE2. To investigate the role of GRP78 in this complex, we knocked down GRP78 in VeroE6-ACE2 cells. Loss of GRP78 markedly reduced cell surface ACE2 expression and led to activation of markers of the unfolded protein response. Treatment of lung epithelial cells with a humanized monoclonal antibody (hMAb159) selected for its safe clinical profile in preclinical models depleted cell surface GRP78 and reduced cell surface ACE2 expression, as well as SARS-2-S-driven viral entry and SARS-CoV-2 infection in vitro. Our data suggest that GRP78 is an important host auxiliary factor for SARS-CoV-2 entry and infection and a potential target to combat this novel pathogen and other viruses that utilize GRP78 in combination therapy.
Highlights
The severe acute respiratory syndrome coronavirus 2 (SARSCoV-2), the causative agent of the COVID-19 global pandemic, utilizes the host receptor angiotensin-converting enzyme 2 (ACE2) for viral entry
Utilizing the dominant negative mutant G227D unable to bind ATP, the T453D mutant unable to bind protein substrates and the R197H mutant, which renders 78-kDa glucose-regulated protein (GRP78) unable to associate with cochaperone DnaJ proteins [20], we probed whether any of these activities is required for GRP78 binding to SARS-2-S and ACE2
Mutant did not, whereas both G227D and T453D mutants were unable to bind ACE2 (Fig. 1D). These results indicate that GRP78 can directly bind to the receptor-binding domain (RBD) of SARS-2-S and the SBD of GRP78 is most critical for interaction between GRP78 and SARS-2-S providing experimental evidence consistent with a previous in silico molecular docking study [4]
Summary
The severe acute respiratory syndrome coronavirus 2 (SARSCoV-2), the causative agent of the COVID-19 global pandemic, utilizes the host receptor angiotensin-converting enzyme 2 (ACE2) for viral entry. SARS-CoV-2-Spike protein (SARS-2-S) responsible for viral attachment and fusion to the host cells exploits angiotensinconverting enzyme 2 (ACE2) as the cellular receptor for viral entry, evidence is emerging that other host factors may serve as critical entry cofactors for productive infection [1, 2].
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