The leukocyte integrin LFA-1 is critically involved in antigen-specific activation of T cells and B cells and in lymphocyte recirculation. In addition to its function as an adhesion receptor, LFA-1 appears to have a role in enhancing T cell activation, and T cell activation modulates the adhesion function of LFA-1. The chief aims of this project are (I) to characterize the equilibrium and kinetic properties of the LFA-1-ICAM-1 interaction, (II) to examine the function of ICAM-1 expression on T cells with respect to recognition of antigen and T cell activation, and (III) to develop reagents, especially monoclonal antibodies, for the study in murine antigen-specific systems, of other LFA-1 ligands. (I) The molecular basis for the modulation of LFA-1 adhesion during activation is unknown, but a possible mechanism is a transition to a conformation with an increased affinity for the LFA-1 counter-receptor, ICAM-1. The equilibrium and kinetic properties of the LFA-1-ICAM-1 interaction on resting and activated lymphocytes will be determined by using a soluble recombinant form of the ICAM-1 molecule in competitive binding assays in which ICAM-1 competes with radiolabeled Fab fragments of anti-LFA-1 monoclonal antibodies, or with radiolabeled ICAM-1, for binding to LFA-1 on cells. This approach has been adopted because of results from preliminary measurements that indicate that the affinity is low. The equilibrium and kinetic measurements will be used to determine (1) if LFA-1 on resting and activated lymphocytes differs in binding to ICAM-1 and (2) if parameters that affect adhesion, such as temperature, divalent metal ion concentration and presence of metabolic inhibitors, have direct effects on the LFA-1-ICAM-1 interaction or affect post-receptor events. (II) To examine the significance of ICAM-1 expression on T cells, cDNAs for the alpha and beta chains on LFA-1 and Mac-1 will be transfected into fibroblast cell lines that have been previously transfected with class II MHC genes. These fibroblast antigen- processing cell lines that express LFA-1, Mac-1, or both integrins will then be analyzed in functional assays for their ability to activate antigen-specific T cell lines. To determine their efficacy in activation of unprimed naive T cells the ability of the integrin-expressing fibroblast lines to activate T cells from T cell receptor transgenic mice will be analyzed. The binding of soluble ICAM-1 to these cell lines will be measured and these data, along with the functional data, will be examined for evidence that LFA-1 and Mac-1 act in a concerted fashion. (III) To determine the role of other LFA-1 ligands in antigen recognition, monoclonal antibodies against murine homologs of recently identified human FLA-1 counter receptors will be derived by immunizing hamsters with purified human proteins and selecting clones that cross-react on mouse. These reagents will be characterized and used in expression and functional studies using antigen-specific T cell clones and T cells from TCR transgenic mice.
| Torigoe, T; Millan, J A; Chan, K W et al. (1994) Protein tyrosine kinase p56-Lck regulates lymphocyte function-associated 1 adhesion molecule expression, granule exocytosis, and cytolytic effector function in a cloned T cell. J Exp Med 180:1115-27 |
| Ybarrondo, B; O'Rourke, A M; Brian, A A et al. (1994) Contribution of lymphocyte function-associated-1/intercellular adhesion molecule-1 binding to the adhesion/signaling cascade of cytotoxic T lymphocyte activation. J Exp Med 179:359-63 |
| Lollo, B A; Chan, K W; Hanson, E M et al. (1993) Direct evidence for two affinity states for lymphocyte function-associated antigen 1 on activated T cells. J Biol Chem 268:21693-700 |
