Purification of the 110-kilodalton glycoprotein receptor for mouse hepatitis virus (MHV)-A59 from mouse liver and identification of a nonfunctional, homologous protein in MHV-resistant SJL/J mice

Abstract

The receptor for mouse hepatitis virus strain A59 (MHV-A59) is a 110to 120-kilodalton (kDa) glycoprotein which is expressed in MHV-susceptible mouse strains on the membranes of hepatocytes, intestinal epithelial cells, and macrophages. SJL/J mice, which are highly resistant to MHV-A59, were previously shown to lack detectable levels of receptor by using either solid-phase virus receptor assays or binding of a monoclonal anti-receptor antibody (MAb) which blocks infection of MHV-susceptible mouse cells. This MAb was used for affinity purification of the receptor glycoprotein from livers of MHV-susceptible Swiss Webster mice. The MHV receptor and an antigenically related protein of 48 to 58 kDa were copurified and then separated by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The first 15 amino acids of the receptor were sequenced, and a synthetic peptide of this amino acid sequence was prepared. Rabbit antiserum made against this peptide bound to the MHV receptor glycoprotein and the 48to 58-kDa protein from livers of MHVsusceptible BALB/c mice and Swiss Webster mice and from the intestinal brush border of BALB/c mice. In immunoblots of intestinal brush border and hepatocyte membranes of MHV-resistant SJL/J mice, the antibody against the amino terminus of the receptor identified proteins that are 5 to 10 kDA smaller than the MHV receptor and the 48to 58-kDa related protein from Swiss Webster or BALB/c mice. Thus, SJL/J mice express a protein which shares some sequence homology with the MHV receptor but which lacks virus-binding activity and is not recognized by the blocking anti-receptor MAb. These results suggest that resistance of SJL/J mice to MIHV-A59 may be due to absence or mutation of the virus-binding domain in the nonfunctional receptor homolog in SJL/J mice. Mouse hepatitis virus (MHV) is a murine coronavirus which causes inapparent enteric or respiratory infection, diarrhea, hepatitis, and acute and chronic demyelinating diseases of the central nervous system (33). Classic studies by Bang and co-workers showed that mouse strains differ markedly in their susceptibility to MHV and that mouse strain susceptibility correlates with permissiveness for MHV replication in peritoneal macrophages cultured from the different mouse strains (2). A single recessive gene, hv-1, was associated with resistance of C3H mice to MHV-2 (34). A wide variety of mouse strains has been tested for susceptibility to several of the many strains of MHV which differ in virulence and tissue tropism (1, 3, 8-10, 16, 18, 19, 26, 27). SJL/J mice and/or peritoneal macrophages cultured from them are highly resistant to infection with the neurotropic strain MHV-JHM and the hepatotropic strain MHV-A59. This resistance trait has been mapped to a single recessive gene at the hv-2 locus near the svp-2 and albino loci on mouse chromosome 7 (15-17, 26). A solid-phase virusbinding assay was used to show that a receptor for MHV-A59 is present on the brush border membranes of enterocytes and hepatocytes from MHV-susceptible...

Amino acid substitutions and an insertion in the spike glycoprotein extend the host range of the murine coronavirus MHV-A59

“…presence of the anti-receptor MAb-CC1 that binds to an epitope on mCEACAM1a that overlaps, but is not identical to, the virus binding site (Chen and Baric, 1996;Schickli et al, 1997;Wessner et al, 1998). MAb-CC1 binds to mCEA-CAM1a with extremely high affinity and prevents MHV infection in vitro and in vivo (Smith et al, 1991;Williams et al, 1990). To further analyze the interactions of the recombinant viruses with mCEACAM1a, we examined the yields of the recombinant viruses from 17 Cl 1 cells treated with MAb-CC1.…”

“…A mouse monoclonal antibody (MAb) to the MHV nucleocapsid protein (N) (anti-N MAb) was kindly provided by Julian Leibowitz (Department of Pathology and Laboratory Medicine, Texas A&M University, College Station, TX). Mouse anti-mCEACAM1a MAb-CC1 and polyclonal rabbit anti-CEACAM1a 649 serum block MHV binding and infection of murine cells (Dveksler et al, 1991;Williams et al, 1990). A mouse MAb directed against the h subunit of cholera toxin (MAb-Ctrl) was used as an isotype-matched control for anti-N MAb and MAb-CC1.…”

Microheterogeneity of S-glycoprotein of mouse hepatitis virus temperature-sensitive mutants

“…The S peplomer protein of MHV is involved in cell to cell fusion, induction of neutralizing antibodies, attachment to the MHV cellular receptor and is a target of cell-mediated immune responses (Collins et al, 1982;Holmes et al, 1981;Wege et al, 1988;Wysocka et al, 1989;Williams et al, 1990). It is an important determinant of cell and organ tropism of MHV and may have a key role in the evolution of persistent infections.…”

B-Cell-Mediated Lysis of Cells Infected with the Neurotropic JHM Strain of Mouse Hepatitis Virus

“…Cells were washed with PBS containing 0.1% bovine serum albumin prior to staining of B cells with fluorescein isothiocyanate-conjugated anti-CD19 (mAb ID3; PharMingen) at 4°C for 15 to 20 min. MHV-R expression was examined by incubation with biotinylated mAb CC-1, which detects CEACAM1a but not CEACAM1b (Williams et al, 1990), and visualized with phycoerythrin-conjugated streptavidin (PharMingen). Samples were analyzed on a FACStar flow cytometer (Becton-Dickinson, Mountain View, CA).…”