Identification, by protein sequencing and gene transfection, of sgp-60 as the murine homologue of CD48.

Abstract

We have recently described a murine lymphocyte protein, provisionally termed sgp-60, which is expressed on virtually all lymphocytes of both T-and B-cell origin. A hamster monoclonal antibody to sgp-60 can inhibit interleukin-2 (IL-2) production, IL-2-receptor expression, and T-cell proliferation, events normally observed after stimulation of T cells with an antibody to the T-cell receptor/CD3 complex or with the lectin concanavalin A. Our previous studies did not reveal the molecular nature of the sgp-60 antigen. Purification of sgp-60 and protein sequencing demonstrate that sgp-60 is identical to the CD48 antigen, a ligand for the CD2 antigen, which is also called Blast-i in humans, BCM1 in mice, and OX-45 in rats. The identity of sgp-60 and CD48 was independently confirmed in gene transfection experiments. The antisgp-60 monoclonal antibody was selectively reactive with COS-7 ceUs transfected with a BCM1 cDNA clone but not with Kb-transfected controls. The results of the present report, together with our previous functional studies, may have implications for the role of CD2 and CD48 in murine T-cell activation.Antigen-specific CD4+ T inducer cells are critically involved in the generation of delayed-type hypersensitivity, antibody, and cytotoxic responses. Normally, CD4+ T cells are in a resting state. Most immune responses therefore depend on the prior activation of CD4+ T lymphocytes, their rapid proliferative expansion, and the lymphokines which they secrete (1).The activation of CD4+ T cells depends critically on cognate interactions of the T cell with an antigen-presenting cell (APC). Physiologic activation is imparted by a heterodimeric T-cell receptor (TCR) with specificity for antigenic peptides complexed to major histocompatibility complex (MHC) proteins. The TCR is associated with a set of nonvariable proteins, collectively termed CD3 (T3) (2). Besides the polypeptides forming the TCR/CD3 complex, a number of molecules participate in the T-cell activation process. Some function as cell-cell interaction molecules and some as signal-transducing elements, and some can serve both functions (2).We have recently described a murine glycoprotein, provisionally termed sgp-60, which is expressed on virtually all lymphocytes of both T-and B-cell origin (3-5). Antibody inhibition studies suggest a functional role for sgp-60 in CD4+ T lymphocytes: addition of the anti-sgp-60 monoclonal antibody (mAb) 5-8A10 or of its Fab fragment significantly inhibits the activation of purified T cells by the lectin Con A or by a mitogenic anti-CD3 mAb (3, 4). Proliferation, interleukin 2 (IL-2) production, IL-2 receptor expression, and generation of second messengers are all affected by the anti-sgp-60 mAb (3,4,6). Only a few antibodies affect T-cell function. Our experiments therefore suggest a role for sgp-60 in T-cell activation.Our previous studies did not reveal the molecular identity of the sgp-60 antigen. In the present report, we close this gap. Partial amino acid sequence of the sgp-60 antigen and gene transfe...

Enhanced murine CD4+ T cell responses induced by the CD2 ligand CD48

“…We have previously employed this system to demonstrate the co-stimulatory activity of murine CD80 [21]. To generate CD48-expressing CHO cells, we cotransfected a full-length CD48 cDNA/pCDM8 clone [22] together with a puromycin resistance plasmid into CHO cells which stably express the I-A d molecule (CHO-I-A d cells) [21]. Following transfection, the cells were drug-selected and the resistant cells sorted for CD48 expression by flow cytometry.…”

“…The CHO-I-A d and CHO-I-A d /CD80 cells have been previously described [21]. CHO-I-A d / CD48 cells were generated by cotransfecting CHO-I-A d cells with a full-length murine CD48 clone [22] and a plasmid encoding puromycin resistance. Transfectants were selected in medium containing 400 ?…”

CD48-deficient mice have a pronounced defect in CD4+T cell activation

“…We initially isolated CD48 genomic clones from a 129 genomic DASH II library, using the CD48 cDNA insert that previously had been isolated (20). A schematic map of the murine Cd48 gene is illustrated in Fig.…”

“…We initially isolated CD48 genomic clones from a 129 genomic DASH II library, using the CD48 cDNA insert that previously had been isolated (20). Sequencing of overlapping genomic clones revealed four distinct exons that correspond to the postulated leader peptide, IgV-like domain, IgC-like domain, and anchor region (J.G.-C., G.J.F., and H.R., unpublished work).…”

Recent advances in graft‐versus‐host disease (GVHD) prevention

“…The in vivo administration of anti-CD2 mAb has been found to inhibit both CD4* and CD8* T cells in a tumor model, presumably affecting the cytotoxic T lymphocytes (CTL) by eliminating T-cell helper function (59,63). Parallel approaches with anti-CD48 mAb have been shown to inhibit IL-2 production, IL-2R expression, IL-4 production (64), and T-cell prohferation (65). The combination therapy of anti-CD2 plus anti-CD48 mAb has synergistic inhibitory effects on alloresponses resulting in tbe indefmite survival of cardiac allografts (64).…”