Cellular entry of the SARS coronavirus

Abstract

Enveloped viruses have evolved membrane glycoproteins (GPs) that mediate entry into host cells. These proteins are important targets for antiviral therapies and vaccines. Several efforts to understand and combat infection by severe acute respiratory syndrome coronavirus (SARS-CoV) have therefore focused on the viral GP, known as spike (S). In a short period of time, important aspects of SARS-CoV S-protein function were unraveled. The identification of angiotensin-converting enzyme 2 (ACE2) as a receptor for SARS-CoV provided an insight into viral tropism and pathogenesis, whereas mapping of functional domains in the S-protein enabled inhibitors to be generated. Vaccines designed on the basis of SARS-CoV S-protein were shown to be effective in animals and consequently are attractive candidates for vaccine trials in humans. Here, we discuss how SARS-CoV S facilitates viral entry into target cells and illustrate current approaches that are used to inhibit this process.

Figure 1

Structural features of the coronavirus (CoV) spike (S)-protein. The localization of the S-gene within the SARS-CoV genome is shown in the upper panel , . S-proteins of coronaviruses exhibit a domain organization that is characteristic of class I fusion proteins. Prototype members of this group of proteins include hemagglutinin (HA) of influenza virus and the gp160 envelope protein of human immunodeficiency virus (HIV) , which are also shown. Generally, these viral glycoproteins can be subdivided into the N-terminal receptor-binding S1 subunit, followed by the S2 subunit containing structural elements required for membrane fusion: a fusogenic peptide and two amino acid stretches with helical symmetry (HR1 and HR2) , . Proteolytic cleavage into the S1 and S2 subunits by host-cell proteases is indicated by a triangular arrow. Functional regions within the NL63 S-protein were predicted using sequence similarity to the closest related coronavirus member hCoV-229E . Abbreviations: AIBV, avian infectious bronchitis virus; hCoV, human CoV; HR, heptad repeat; MHV, murine hepatitis virus; SARS, severe acute respiratory syndrome.

Figure 2

Routes of virus entry into target cells. Binding of a viral glycoprotein (GP) to a receptor can induce conformational alterations, which facilitate fusion of the viral and plasma membrane leading to release of the viral nucleocapsid into the cytoplasm (left). Alternatively, binding to the receptor can be followed by uptake of the virion into an endosomal compartment (right) , . Proton influx into the endosome can then trigger the membrane fusion activity in GP. Inhibitors of acidification, such as ammonium chloride (NH₄Cl) or bafilomycin A (BAF), can be used to dissect both pathways because they specifically block pH-dependent membrane fusion. For entry of SARS-CoV S-bearing particles, the endosomal pathway appears to be important , , . However, an acid environment has been shown to be dispensable for the fusion of cells that express the S-protein to adjoining cells that express the SARS-CoV receptor ACE2 (angiotensin-converting enzyme 2) , .

Figure 3

Membrane fusion and its inhibition. Binding of SARS-CoV spike (S)-protein to ACE2 (angiotensin-converting enzyme 2) promotes internalization into endosomes, where the low pH environment triggers fusion activity in the S-protein. Membrane fusion is driven by the S2 subunit of S, which contains a putative fusion peptide and two heptad repeats (HRs) , . Upon insertion of the fusion peptide into the host cell membrane, HR1 folds back onto HR2, forming a highly stable six-helix bundle , , , . The formation of the six helix bundle brings the viral and the host cell membrane in close proximity and ultimately promotes membrane fusion (upper panel). The HRs are attractive targets for antiviral therapy; synthetic peptides mimicking HR2 can complex the HR1 regions, which inhibits the formation of the six-helix bundle (lower panel) , , .