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We studied the role of CD43 (leukosialin/sialophorin), the negatively charged sialoglycoprotein of leukocytes, in the binding of mycobacteria to host cells. CD43-transfected HeLa cells bound Mycobacterium avium, but not Salmonella typhimurium or Shigella flexneri. Quantitative bacteriology showed that macrophages (M(phi)) from wild-type mice (CD43(+/+)) bound M. avium, Mycobacterium bovis (bacillus Calmette-Guérin), and Mycobacterium tuberculosis (strain H37Rv), whereas M(phi) from CD43 knockout mice (CD43(-/)-) did not. Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43(+/+) M(phi). The inability of CD43(-/)- M(phi) to bind M. avium could be restored by addition of galactoglycoprotein (Galgp), the extracellular mucin portion of CD43. The effect of Galgp is not due to opsonization of the bacteria, but required its interaction with the M(phi) other mucins had no effect. CD43 expression by the M(phi) was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-alpha production, which likewise could be reconstituted by Galgp. In contrast, interleukin (IL)-10 production by M. avium-infected M(phi) was CD43 independent, demonstrating discordant regulation of TNF-alpha and IL-10. These findings describe a novel role of CD43 in promoting stable interaction of mycobacteria with receptors on the M(phi) enabling the cells to respond specifically with TNF-alpha production.

The mAb 1B11 has been characterized as recognizing the activation-associated glycoform of murine CD43, a heavily O-glycosylated protein implicated in leukocyte homing. When hemopoietic cells from CD43-/- mice were stained with 1B11, CD43-independent binding of 1B11 was observed on peripheral CD8 T cells and at low levels on thymocytes, while no binding was detected on CD4 T cells, B cells, or bone marrow cells. Levels of 1B11 staining were comparable in lymph node CD8+ T cells from both CD43-/- mice and CD43+/+ mice. We sought to identify the CD43-independent target of 1B11 expressed on CD8 T cells. Previous work had demonstrated that neuraminidase treatment of lymph node cells (LNC) enhanced 1B11 binding on CD43+/+ LNC; this enhancement was also observed in CD43-/- LNC. We show that neuraminidase-enhanced 1B11 binding in CD43-/- LNC and EL4 thymoma cells is CD43 independent and that 1B11 detects a novel target of apparent mass of approximately 200 kDa identified as a hyposialylated form of CD45RB preferentially expressed on peripheral CD8, but not CD4, T cells. Our data also show that the recognition of CD43 and CD45RB by 1B11 is differentially affected by O-linked glycosylation and sialic acid. Whereas 1B11 recognition of CD43 on activated T cells required both core 2 O-glycan branching and sialic acid, 1B11 recognition of CD45 only occurred in the absence of both core 2 glycosylation and sialic acid.

CD43, a large highly glycosylated molecule, is arguably the most abundant molecule on the surface of T cells. Nevertheless, the function of CD43 remains unclear. Utilizing fluorescence microscopy, we find that CD43 is excluded from the T cell-APC contact site. This exclusion is Ag dependent since optimal CD43 exclusion requires Ag-pulsed APC, and since signaling through CD3, in the absence of any other receptor ligand interactions, can induce the modulation of CD43. These data suggest that CD43 may function as a barrier to nonspecific T cell-APC interactions that is removed as a result of T cell activation. Exclusion from the interaction site is a unique feature of CD43 and not universally found for all large highly glycosylated molecules since CD45 is not excluded. Thus, CD43 may represent a novel regulatory molecule on the T cell surface that can direct T cell interactions by changing its location on the cell surface.

Accessory molecule interactions can be critical in determining the outcome of a T cell's encounter with antigen. Cell adhesion proteins may augment T cell responses by facilitating TCR engagement of the antigen-MHC complex, while co-stimulatory molecules may deliver distinct signals that modulate T cell responsiveness. CD43 (leukosialin, sialophorin) has been suggested to influence cell activation by steric hindrance based upon the large size and glycosylation of the protein, as well as the relative abundance of the protein on the cell surface. In this paper we examine both in vitro and in vivo T cell-dependent responses in CD43-deficient mice. We demonstrate that T cells from CD43-deficient mice are hyper-responsive following both in vivo and in vitro activation, and that this is observed in response to not only TCR-CD3-mediated stimulation, but also following receptor-independent activation. This data suggests that mechanisms other than non-specific steric hindrance are important in the regulation of T cell activation by CD43.

We report that the cell surface mucin CD43 acts as an anti-adhesin on T lymphocytes. CD43-deficient murine lymphocytes homed significantly more frequently to secondary lymphoid organs than wild-type cells. Intravital microscopy of peripheral lymph node venules revealed that CD43-deficient lymphocytes were twice as likely to tether, roll, and stick than wild-type cells. This effect was due to CD43 interference with the homing receptor, L-selectin, and was most pronounced in venules with low L-selectin ligand density. In vitro, CD43-deficient cells tethered to L-selectin ligands more efficiently and rolled more slowly than wild-type lymphocytes. Thus, CD43 exerts a negative regulatory effect on T cell trafficking by counterbalancing L-selectin-mediated adhesion.

The leukocyte sialyloglycoprotein CD43 exhibits features of a signal transducing molecule and is thought to be important for T-cell activation and adhesion. However, cellular biochemical events in which CD43 participates remain poorly understood. Here we provide evidence that CD43 regulates tyrosine phosphorylation of a specific substrate in T cells. A 93-kD tyrosine phosphoprotein was identified specifically in the CD43+ T-cell line CEM, but not in their CD43-deficient counterparts derived by gene targeting. The 93-kD phosphoprotein was detected in the CD43-deficient CEM cells after transfection with CD43 cDNA, and it could be specifically phosphorylated in lysates from the CD43-deficient cells by incubation with a CD43 immunoprecipitate obtained from the CD43+ cells. Expression of CD43 in HeLa cell transfectants was associated with the appearance of novel phosphoproteins including one with a molecular weight of approximately 93 kD, confirming that tyrosine phosphorylation of cellular substrates results specifically from CD43 expression. We conclude that CD43 regulates tyrosine phosphorylation of a 93-kD T-cell substrate.

CD43 is a cell-surface sialoglycoprotein expressed by a variety of haematopoietically derived cells, including T lymphocytes. Earlier observations of defective CD43 expression by T lymphocytes from boys with the X-chromosome-linked Wiskott-Aldrich syndrome suggested the importance of CD43 in lymphocyte function. Subsequent studies have suggested that CD43 facilitates leukocyte adhesion and has a co-stimulatory role during T-cell activation. To define the physiologically relevant function(s) of CD43, we have generated CD43-knockout mice. We report here that CD43-deficient T cells from such mice show a marked increase in their in vitro proliferative response to concanavalin A, anti-CD3, the superantigen SEB and allostimulation. Additionally, CD43-deficient T cells show a substantial enhancement in homotypic adhesion and in their ability to bind different ligands, including fibronectin and the intercellular adhesion molecule ICAM-1. Vaccinia-virus-infected CD43-knockout mice mounted an augmented anti-vaccinia cytotoxic T-cell response compared with their wild-type littermates, yet developed an increased virus load. We conclude that CD43 negatively regulates T-cell activation and adhesion and is important for viral clearance.

CD43 is a cell surface sialoglycoprotein expressed by most cells of hematopoietic origin, including all T lymphocytes. Elimination of CD43 expression by gene targeting in the CEM T cell line results in its increased homotypic adhesion and binding to HIV-1 gp120. Here we report that the CD43-negative CEM cells show increased susceptibility to HIV-1 infection and increased viral replication compared with the parental CD43+ CEM cell line. Increased HIV-1 replication also was observed in CEM cells with diminished CD43 expression secondary to functional inactivation of a single CD43 allele. The CD43- CEM cells were more susceptible to HIV-1-induced cytopathicity than their CD43+ counterparts. HIV-1 replication also was increased in the CD43- CEM cells after transfection with the infectious HIV molecular clone pNL4-3. These data suggest that factors that diminish CD43 expression on T lymphocytes may enhance their susceptibility to HIV-1 infection.

CD43 is a major membrane sialoglycoprotein expressed by cells of hematopoietic origin. One property of CD43 is its ability to interfere with heterotypic and homotypic cellular adhesion. To determine whether CD43 expression can affect cell functions requiring intercellular adhesion, we compared a CD43-positive human T cell line (CEM) and its CD43-negative counterpart derived by gene targeting for susceptibility to cell-mediated lysis. CD43-negative CEM cells were more susceptible than CD43-positive cells to lysis by allospecific T cell lines derived from several donors. Induction of CD43 expression on transfected HeLa cells also imparted resistance to lectin-mediated lysis by a CD8+ T cell clone. The effect of CD43 expression on reducing susceptibility to lysis was more pronounced in short-term cytotoxicity assays and tended to disappear as the time of contact between the effector cell and its target increased. The enhanced susceptibility of CD43-negative cells to lysis was not associated with increased expression of adhesion molecules known to mediate antigen-independent cellular adhesion. Sialic acid residues on CD43 contributed to the CD43 protective effect. These results suggest that either diminished CD43 expression or incomplete sialylation may render hematopoietic cells more susceptible to T lymphocyte-mediated cytolysis.

CD43 is a major leukocyte surface glycoprotein thought to have important functions for T lymphocyte adhesion and activation. We investigated the function of CD43 by using gene targeting to eliminate CD43 expression in the human T lymphocyte line CEM and then testing their adhesive phenotype. Loss of CD43 expression by the CEM cells enhanced their homotypic adhesion and binding to two distinct ligands, fibronectin and HIV-1 gp120. The enhanced homotypic adhesion was blocked specifically by an anti-beta 1 integrin mAb, and the enhanced binding to fibronectin and gp120 was blocked specifically by anti-beta 1 integrin and anti-CD4 mAb, respectively. Partial reconstitution of CD43 expression in the CD43-negative cells resulted in a corresponding reversion to a less adhesive phenotype. These data suggest that CD43 interferes with T lymphocyte adhesion and that CD43 can regulate lymphocyte adhesion by providing a threshold that must be overcome for cell-cell and cell-ligand interactions to occur.

CD43 is a cell-surface sialoglycoprotein of uncertain physiologic function expressed to various degrees by most leukocytes. We tested whether or not CD43 participates in intercellular adhesion by comparing the binding of human T lymphocytes to transfected HeLa cells stably expressing CD43 and sham-transfected HeLa cells (CD43-negative). Significantly fewer T lymphocytes adhered to the CD43-positive HeLa cells than to the CD43-negative HeLa cells. Diminished T-cell adherence to the CD43-positive HeLa cells was seen for all T lymphocytes tested, irrespective of their source or derivation. Antibody-blocking experiments revealed that CD43 interference with T-cell adhesion largely represented interference with T-cell leukocyte function-associated antigen 1 binding to HeLa cell intercellular adhesion molecule 1. The CD43 anti-adhesion effect was not overcome by treating cells with phorbol 12-myristate 13-acetate, a chemical that increases the binding avidity of leukocyte function-associated antigen 1 for intercellular adhesion molecule 1. However, neuraminidase treatment of the HeLa cell transfectants diminished the CD43 antiadhesion effect. These data indicate that CD43 expression by opposing cells can interfere with cell-cell adhesion. The data also suggest that CD43 might regulate T-cell adhesion by interfering with leukocyte function-associated 1 binding to intercellular adhesion molecule 1, a major activation-induced adhesion pathway among lymphocytes.

Sera from human immunodeficiency virus type 1 (HIV-1)-infected and -noninfected individuals were screened for antibodies that could bind to native T cell differentiation antigens. Antibodies that could immunoprecipitate CD43 (sialophorin, leukosialin) from a T cell lymphoma line were detected in sera from 27% of patients, and antibodies that could bind specifically to transfected cells expressing CD43 were detected in 47% of patients. The anti-CD43 antibodies were related to HIV-1 infection in that no patients with other chronic viral infections or systemic lupus erythematosus contained such antibodies in their sera. The anti-CD43 autoantibodies bound to a partially sialylated form of CD43 expressed by normal human thymocytes, but not by normal, circulating T lymphocytes. However, the determinant(s) recognized by the anti-CD43 autoantibodies was present on a large proportion of circulating T lymphocytes, but masked from antibody recognition by sialic acid residues. These results demonstrate that HIV-1 infection is specifically associated with the production of autoantibodies that bind to a native T cell surface antigen.