Identification of monoclonal antibody epitopes and critical residues for rhinovirus binding in domain 1 of intercellular adhesion molecule 1.

Abstract

Intercellular adhesion molecule 1 (ICAM-1) is the cellular receptor for the major group of human rhinoviruses (HRVs) and the adhesion ligand of lymphocyte function-associated antigen 1. Analysis of a series of chimeric exchanges between human and murine ICAM-1 shows that two distinct epitopes recognized by monoclonal antibodies that block rhinovirus attachment and cell adhesion map to the N-terminal first domain of ICAM-1. Furthermore the specificity for HRV binding is entirely contained within the first 88 amino acids. Mutagenesis of the four sites of N-linked glycosylation within the second domain shows that carbohydrate is not involved in virus recognition. Homologue replacement mutagenesis localizes the epitopes for virus-blocking antibodies to two regions of domain 1 predicted to form .3 strand D and the loop between the F and G strands of an immunoglobulinfold structure. Analysis of virus binding to the mutants predicts a large surface of contact between HRV and ICAM-1 domain 1 but shows that the regions most important for virus binding are coincident with the monoclonal antibody epitopes.There are more than 100 distinct serotypes of human rhinoviruses (HRVs), the primary causative agent of the common cold (1). Viral entry mediated by binding to a cellular receptor is a critical first step in the infection process and is an important determinant of viral tropism. Ninety percent of HRV serotypes utilize a common cellular receptor to initiate infection (2, 3), which we have recently shown is intercellular adhesion molecule 1, or ICAM-1 (4). ICAM-1 is a member of the IgG supergene family; it interacts with the leukocyte integrin lymphocyte function-associated antigen 1 (LFA-1) (5, 6). The molecule is an integral membrane glycoprotein with an extracellular region of453 amino acids containing five domains with sequence similarity to the IgG constant regions.The three-dimensional structure of HRV14, which binds to ICAM-1, and of HRV1A, which binds to the as yet unidentified minor receptor, has been determined (7,8). Determination of the structure of HRV14 led Rossmann to propose the canyon hypothesis, which suggests that a 20-A-wide surface depression, which encircles the fivefold axis of symmetry of each icosahedral face of the virus, may contain the receptor binding site. Support for the canyon hypothesis comes from site-directed mutagenesis of canyon residues, which alter the receptor binding properties of HRV14 (9), and from studies with capsid binding drugs, which induce a conformational change in the floor of the canyon and prevent receptor binding (10). The dimensions of the canyon are sufficient to accommodate a single unpaired IgG domain, and it has recently been shown by electron microscopy that ICAM-1 and the related neural cell adhesion molecule (NCAM) have long elongated structures consistent with an end-to-end arrangement of unpaired IgG domains (11,12). The N-terminal domain of ICAM-1 is therefore likely to project furthest from the cell surface and be most accessible to virus. Furthermore...

Discrimination among Rhinovirus Serotypes for a Variant ICAM-1 Receptor Molecule

“…Of special interest is Pro70 of ICAM-1 that is stacked on top of Pro3178 in HRV14 or on top of the equivalent Pro3180 in HRV16. When Pro70 is mutated to almost any other amino acid, the binding between receptor and virus is eliminated (32,41). Another hydrophobic residue, Phe3086 in HRV14 (Tyr3086 in HRV3), is close to the Pro70-Pro3178 interaction, which may further enhance the local hydrophobic environment.…”

Regulation of the expression of intercellular adhesion molecule 1 in cultured human endothelial cells derived from rheumatoid synovium

“…The media were removed and the cells fixed for 10 minutes at 23°C with 3.7% formalin in PBS. The cells were washed and then incubated with a 1:1,OOO dilution of the first antibody (anti-ICAM-1 C78.5 [14]; kindly supplied by Dr. J. Greve, Molecular Therapeutics Inc., West Haven, CT) or nonimmune control IgG in PBS-0.1% BSA for 2 hours at 23°C. The cells were washed in PBS-BSA, then blocked for 1 hour in PBS-3% BSA.…”

The Coxsackievirus and Adenovirus Receptor: Glycosylation and the Extracellular D2 Domain Are Not Required for Coxsackievirus B3 Infection

“…ceptor function, although the efficiency of binding and infection was increased by the addition of some, but not all, heterologous immunoglobulin-like domains (18)(19)(20)41).…”

“…However, studies with PV, HRV, and hepatitis A virus (HAV) have shown that deletion of the membrane-proximal extracellular domains decreases virus binding to the receptor, as well as infection of the cells (13,(15)(16)(17). Replacement of these proximal domains with homologous domains from other species restored normal receptor function (18,19), but replacement with heterologous protein domains did not (16,20). Thus, domains that are located membrane proximal to the virus-binding D1 domain are important to maintain virus receptor properties, but the mechanisms that are involved are not well understood.…”

“…CAR has two N-glycosylation sites, one on each of its extracellular domains (N106 in D1 and N201 in D2). Glycosylation of ICAM-1 is not required for rhinovirus binding (19,24) or for poliovirus interaction with PVR (25). In contrast, the binding and infection efficiency of hepatitis A virus (HAV) depends on glycosylation of the HAV cellular receptor havcr-1, and deglycosylation results in decreased susceptibility of the cells to HAV infection (26).…”