Christine B. Cardellichio

B lymphocyte and macrophage expression of carcinoembryonic antigen-related adhesion molecules that serve as receptors for murine coronavirus

The expression of carcinoembryonic antigen (CEA)‐related glycoproteins that have been associated with intercellular adhesion and that serve as receptors for mouse hepatitis virus (MHV) was analyzed in cells from the immune system of BALB/c mice using immunolabeling and RNA polymerase chain reaction amplification of receptor transcripts. These glycoproteins, which are called biliary glycoproteins, were highly expressed in B lymphocytes, including cells of the B‐la (CD5+) lineage, and in macrophages, but were not detectable in resting T lymphocytes. Similarly, murine cell lines of B cell and macrophage origin expressed messenger RNA encoding CEA‐related molecules, while the corresponding mRNA was only slightly detectable in a T cell line. These CEA‐related cell adhesion glycoproteins were also expressed in endothelial cells. Therefore, their specific interaction with their so far unknown ligand may be of functional importance in cellular interactions in the immune response.

Monoclonal Antibody to the Receptor for Murine Coronavirus MHV-A59 Inhibits Viral Replication In Vivo

Abstract Because many strains of mouse hepatitis virus (MHV) infect laboratory mice, no effective vaccine has yet been developed. An alternative approach to control MHV disease is the use of a host cell receptor-targeted ligand. Toaddress the potential usefulness of this approach, a monoclonal antibody directed against the host cell receptor for the coronavirus MHV-A59 was administered to infant mice that were then challenged oronasally with 104 intracerebral infant mouse median lethal doses of MHV-A59. Antibody treatment of virus-challenged mice resulted in lower proportions of mice with MHV-A59 in target organs and markedly reduced viral titers in these organs compared with mock-treated infected mice. Some antibody-treated infected mice survived for 7 days after viral challenge, whereas no mock-treated, infected mice survived beyond day 3 after viral inoculation. These results support a receptor-targeted approach to intervention in coronavirus disease.

Multiple receptor-dependent steps determine the species specificity of HCV-229E infection.

Human coronavirus (HCV)-229E causes disease only in humans and grows in human cells and in cells of other species that express recombinant human aminopeptidase N (hAPN), the receptor for HCV-229E. We compared the species specificity of HCV-229E infection with the species specificity of virus binding using immunofluorescence, assay of virus yields, fluorescence activated cell sorting and a monoclonal antibody directed against hAPN that blocks infection. We found that HCV-229E binds to intestinal brush border membranes (BBM) and to membranes of cell lines from cats, dogs, pigs, and humans, however the virus only infects two of these species. HCV-229E will not bind to BBM or to membranes from cell lines derived from hamster or mice. Animal coronaviruses related to HCV-229E, including FIPV, CCV, and TGEV bind to cell membranes from cats, dogs, cows, pigs and humans (but not mice), while each virus infects cells from only a subset of these species. Infectious genomic HCV-229E RNA, can infect cells of all of these species. These data suggest that the species-specificity of infection for this serogroup of coronaviruses is determined at the levels of virus binding and penetration. Since binding of viral spike glycoprotein to cellular receptors is not the only limiting factor, we suggest that one or more steps associated with virus penetration may determine the species specificity of infection with the HCV-229E serogroup of coronaviruses.

Characterization of a New Gene that Encodes a Functional MHV Receptor and Progress in the Identification of the Virus-Binding Site(s)

Several splice variants of the murine biliary glycoprotein 1 (Bgp 1) gene in the carcinoembryonic antigen gene superfamily serve as cellular receptors for mouse hepatitis virus. RNAPCR and immunoblot analysis of the receptor in inbred mouse strains showed that the glycoproteins expressed in SJL/J mice are encoded by an allelic variant of the Bgp 1 gene, named Bgp 1b. We recently cloned and characterized a second gene, Bgp 2, that encodes a functional MHV receptor glycoprotein which is not recognized by anti-MHVR MAb-CC1. A third gene related to Bgp 1 was cloned and expressed and shown to encode a soluble protein called Cea-10 that differs significantly in its N-terminal domain from Bgp 1 and Bgp 2. Chimeric proteins constructed between the different murine Bgps and point mutations in the prototype MHV receptor, Bgp 1a or MHVR, were analyzed to further characterize the MAb-CC1-binding and virus-binding domains within the N terminal domain of the receptor. Thus, the murine host for MHV expresses multiple splice variants of mRNAs encoded by several different Bgp-related genes which differ in their ability to serve as MHV receptors. The differential expression of these genes in different murine tissues may help to explain the tissue tropism of MHV strains.

Expression of MHV-A59 Receptor Glycoproteins in Susceptible and Resistant Strains of Mice

Band and Warwick showed that inbred mouse strains differed in their susceptibility to virulent strain of mouse hepatitis virus, MHV-2, and that peritoneal macrophages cultured from these mouse strains reflected their differences in susceptibility to the virus (1). Adult SJL/J mice are highly resistant to MHV-JHM and MHV-A59 (2,12,13). Our laboratory showed that membranes from the liver and intestinal epithelial cells of BALB/c mice could bind MHV-A59 virions in a solid phase assay using undenatured membrane proteins, whereas membranes from SJL/J mice did not bind virus, suggesting that differences in binding of virus to cells from different mouse strains could account for their differences in susceptibility to MHV (3). In virus-overlay protein blot assays of intestinal brush border and liver membrane proteins, MHV-A59 virions bound strongly to a 58 kDa glycoprotein from BALB/c mice, but the virus did not recognize any proteins from SJL/J mouse tissues (3,4). Anti-receptor monoclonal antibody MAb-CC1 that blocked infection of murine cell lines with MHV-A59 recognized the 110–120 kDa and 58 kDa membrane glycoproteins of BALB/c mice, but no proteins from SJL/J membranes (6,7).

Is the 110K Glycoprotein the Only Receptor for MHV and Does Its Expression Determine Species Specificity

Coronaviruses exhibit strong tissue tropisms and species specificities, and the molecular mechanisms for these tropisms are of considerable interest. For mouse hepatitis virus, strain A59 (MHV-A59), a solid phase assay was developed to detect binding of virions to plasma membranes from normal target tissues of susceptible mice (1). Using a virus overlay protein blot assay, MHV-A59 was shown to bind specifically to a 100 to 110K protein from liver or intestine membranes of MHV-susceptible BALB/c mice. The specificity of virus binding was demonstrated by the observations that other enterotropic murine viruses did not bind to the same membrane protein and that MHV-A59 did not bind to any proteins from intestine or hepatocyte membranes from SJL/J mice, which are highly resistant to infection with MHV-A59 (2,3). Thus, SJL/J mice may be resistant to infection with MHV-A59 because the virus fails to bind to its normal target tissues, possibly because the virus-binding moiety is absent from the SJL/J plasma membranes.