REGULATION OF VLA INTEGRIN FUNCTION ON T-CELLS

Molecules which mediate T cell adhesion to other cells and proteins are critical to T cell migration and recognition. The functional activity of adhesion molecules must be carefully regulated so that T cells can alternate appropriately between adhesive and nonadhesive states. The broad objective of this application is to understand the regulatory mechanisms that govern the function of T cell adhesion molecules. The specific focus is on the VLA integrins VLA-4, VLA-5, and VLA-6, which mediate cell adhesion to the extracellular matrix proteins fibronectin (VLA-4 and VLA-5) and laminin (VLA-6). VLA function on resting T cells is regulated by receptor-mediated T cell activation in an undefined manner: CD3- and CD2- mediated activation induces a rapid induction of strong adhesion to fibronectin (FN) and laminin (LN) without a change in quantitative levels of VLA integrin expression. This application will characterize the cellular responses that regulate activation-dependent VLA function on human T cells. i) Monoclonal antibodies (mAb) will be generated that inhibit the activation-dependent adhesion of resting T cells to LN via VLA-6. These mAb will be used to characterize the costimulatory effects of VLA-6 mAb on CD3- and CD2-mediated T cell proliferation and to identify additional cell surface molecules that regulate VLA-6-dependent adhesion to LN. ii) The interrelationship between activation-dependent T cell binding to FN and three intracellular responses (Ca++ mobilization, changes in organization of the cytoskeletal protein actin, and tyrosine phosphorylation) triggered by receptor-mediated activation will be examined using specific pharmacologic agents which induce or inhibit these activation responses. iii) The Jurkat T cell line, which, like resting T cells, does not bind strongly to FN unless activated, will be used to isolate and characterize somatic mutants that no longer adhere to FN after activation. Mutants will be isolated after gamma-irradiation by activating the irradiated cells and enriching for nonadherent mutants by multiple steps of panning of FN. The defects which result in nonadherence in these mutants will be characterized. These studies will provide important insight into the mechanisms that regulate T cell adhesion and how adhesion molecules contribute to T cell function. These findings will also have important applications for understanding how disregulation of adhesion may contribute to diseased states, and will be beneficial in identification of potential therapeutic approaches for targeting or modulating immune responses by regulating adhesive interactions.