Cooperative Involvement of the S1 and S2 Subunits of the Murine Coronavirus Spike Protein in Receptor Binding and Extended Host Range

Abstract

The entry of enveloped viruses into cells requires attachment of the virion to the host cell, followed by fusion of the viral membrane with a membrane of the target cell. While some viruses contain glycoproteins that mediate both virus-cell attachment and virus-cell fusion, as is the case for coronaviruses, for others, such as paramyxoviruses, these functions are served by different glycoproteins. In both cases, however, the viral fusion proteins undergo dramatic conformational changes upon activation, which are tightly controlled in time and place in order to effect the proper merging of viral and cellular membranes.Coronaviruses are enveloped viruses that contain a large single-stranded RNA genome of positive polarity. Their envelope accommodates three or four membrane proteins of which the membrane (M), envelope (E), and spike (S) proteins are common to all (reviewed by de Haan and Rottier [15]). The S protein is a relatively large, 1160-to 1452-amino-acid-long type I glycoprotein, trimers of which form the petal-shaped projections on the surface of the virion that give rise to the characteristic corona solis-like appearance. The S proteins from some but not all coronaviruses are cleaved as a late step during their maturation (reviewed by Cavanagh [8]), for which a furin-like enzyme was shown to be responsible in the case of mouse hepatitis virus (MHV) strain A59 (16). The resulting aminoterminal S1 subunit and the membrane-anchored S2 subunit remain noncovalently linked. It has been suggested that the S1 subunit constitutes the globular head, while the S2 subunit forms the stalk-like region of the spike (8, 9).The two functions of the coronavirus S protein appear to be spatially separated. The S1 subunit (or the equivalent part in viruses with uncleaved S protein) is responsible for receptor binding, and the S2 subunit is responsible for membrane fusion. For MHV, the receptor-binding domain has been mapped to the amino-terminal 330 residues of the S molecule (23, 42). For transmissible gastroenteritis virus (20), human coronavirus 229E (4, 7), and severe acute respiratory syndrome (SARS) coronavirus (1, 44), the receptor-binding domains have also been mapped to the S1 subunit, though to different regions therein. The ectodomain of the S2 subunit contains two heptad repeat (HR) regions (9) (Fig. 1), characteristic of coiled coils, while the fusion peptide (FP) is predicted to be located amino terminally of the first HR region (HR1) (5). Binding of the S1 subunit to the (soluble) receptor has been shown to trigger conformational changes that supposedly fa-

Synthetic Reconstruction of Zoonotic and Early Human Severe Acute Respiratory Syndrome Coronavirus Isolates That Produce Fatal Disease in Aged Mice

“…Similar adaptations have been shown previously for another icSARS-CoV bearing the GD03 S glycoprotein at posi- tions 7 and 613 (10). Adaptive mutations in the S glycoprotein have been implicated in coronavirus adaptation and extended host studies (9), although involvement of unknown mutations in ORF1a/ORF1b cannot be excluded. The lack of mutations in the S genes of strains icUrbani, icCUHK-W1, and icGZ02 as well as in the rest of the 3Ј end of the SARS-CoV genome indicates that the reverse genetic system can be readily used to introduce mutations in the S gene without subsequent reversion or extensive adaptation, making this an attractive tool for studying the effects of heterogeneity in the S gene in a stable genetic background of the Urbani isolate.…”

“…Three amino acid changes in the S glycoprotein of SARS-CoV were associated with a lethal phenotype in aged mice, including one each in the receptor binding domain, the putative FP region, and the HR-2 region. Mutations in the HR and FP domains of mouse hepatitis virus have previously been shown to affect host ranges in vitro and pathogenesis in vivo (9,59). Mice infected with viruses bearing these three mutations showed clinical signs, lost significant amounts of weight, and died by day 4.…”

Contributions of the S2 spike ectodomain to attachment and host range of infectious bronchitis virus

“…In coronavirus infections, when processes of virus attachment and entry cannot be distinguished from each other, a concerted action between S1 and S2 has been observed. For example, the extended host range of MHV strains often requires a combination of mutations in S1 and S2 (de Haan et al, 2006;Navas-Martin et al, 2005;Saeki et al, 1997;Schickli et al, 2004;Thackray and Holmes, 2004). When elucidated in more detail, the communication between these regions was particularly contributing to virus-cell fusion and spread of progeny virus.…”

“…The S2 domain, but not HS binding site, is required for gain of binding of Beaudette spike to susceptible tissues Currently, all evidence points toward a sole contribution for the viral glycoprotein spike in coronaviral attachment. While the S1 is generally assumed to contain the RBD, the C-terminal S2 domain might act in concert with S1 during entry and account for the extended host range, as was shown for a murine hepatitis virus (MHV) strain with extended tropism (de Haan et al, 2006). In addition, the S2 of Beaudette has a heparin-binding consensus XBBBXXB sequence (where X is a hydropathic amino acid and B is a basic residue, as previously defined by Cardin and Weintraub, 1989).…”

Identification of two novel B cell epitopes on porcine epidemic diarrhea virus spike protein

“…Based on bioinformatics analysis of the S protein structure of other coronaviruses, the S1 domain is thought to form the spherical head of the S protein, and is easily accessible to neutralizing antibodies. The S2 domain forms the stem portion and trans-membrane structure of the S protein, and it seldom induces antibodies because of the inaccessibility of this region in intact virions (Holmes, 2001;Wu et al, 2004;De Haan et al, 2006). The epitopes SS2 and SS6 are presumed to be located in the spherical head of the S protein of PEDV.…”