Mouse hepatitis virus strain A59 and blocking antireceptor monoclonal antibody bind to the N-terminal domain of cellular receptor.

Mouse hepatitis virus (MHV) strain A59 uses as cellular receptors members of the carcinoembryonic antigen family in the immunoglobulin superfamily. Recombinant receptor proteins with deletions of whole or partial immunoglobulin domains were used to identify the regions of receptor glycoprotein recognized by virus and by antireceptor monoclonal antibody CC1, which blocks infection of murine cells. Monoclonal antibody CC1 and MHV-A59 virions bound only to recombinant proteins containing the entire first domain of MHV receptor. To determine which of the proteins could serve as functional virus receptors, receptor-negative hamster cells were transfected with recombinant deletion clones and then challenged with MHV-A59 virions. Receptor activity required the entire N-terminal domain with either the second or the fourth domain and the transmembrane and cytoplasmic domains. Recombinant proteins lacking the first domain or its C-terminal portion did not serve as viral receptors. Thus, like other virus receptors in the immunoglobulin superfamily, including CD4, poliovirus receptor, and intercellular adhesion molecule 1, the N-terminal domain of MHV receptor is recognized by the virus and the blocking monoclonal antibody.

Binding of the coronavirus mouse hepatitis virus A59 to its receptor expressed from a recombinant vaccinia virus depends on posttranslational processing of the receptor glycoprotein.

Recently, we showed that a murine member of the carcinoembryonic antigen family of glycoproteins serves as a cellular receptor (MHVR) for the coronavirus mouse hepatitis virus A59 (MHV-A59) (G. S. Dveksler, M. N. Pensiero, C. B. Cardellichio, R. K. Williams, G.-S. Jiang, K. V. Holmes, and C. W. Dieffenbach, J. Virol. 65:6881-6891, 1991; R. K. Williams, G.-S. Jiang, and K. V. Holmes, Proc. Natl. Acad. Sci. USA 88:5533-5536, 1991). To examine the role of posttranscriptional modification of MHVR on virus-receptor interactions, a vaccinia virus-based expression system was employed. Expression from the vaccinia virus recombinant (Vac-MHVR) in BHK-21 cells resulted in high levels of MHVR glycoprotein on the cell surface and made these cells susceptible to MHV-A59 infection. Nonglycosylated core MHVR proteins were made in Vac-MHVR-infected BHK-21 cells in the presence of tunicamycin by in vitro translation of MHVR mRNA in a rabbit reticulocyte cell-free system in the absence of microsomal membranes and by expression of an N-terminal deletion clone of MHVR lacking its signal peptide. These three nonglycosylated MHVR proteins were recognized by polyclonal antibody against affinity-purified receptor but did not bind antireceptor monoclonal antibody (MAb) CC1 or MHV-A59 virions. Partial glycosylation of MHVR, either expressed in Vac-MHVR-infected cells treated with monensin or synthesized by in vitro translation with microsomal membranes, restored both the MAb CC1- and the virus-binding activities of the MHVR glycoprotein. Deletion of 26 amino acids at the carboxyl terminus of MHVR resulted in a secreted protein which was able to bind MAb CC1 and MHV-A59. These results suggest that either a carbohydrate moiety is an element of the MHVR-binding site(s) for virus and MAb CC1 or a posttranslational membrane-associated process is required for functional conformation of the receptor glycoprotein.