We demonstrated that alpha3 beta1 and alpha6 beta1 integrins are laminin/merosin receptors expressed in human thymocytes. A small percentage (15%) of human thymocytes bind to immobilized laminin, and even fewer bind to merosin, the laminin isoform normally present in the thymus. This binding, however, can be increased to 39 to 41% after activation of thymocytes with Mn2+ (or PMA). Both alpha3 beta1 and alpha6 beta1 were shown to participate in thymocyte binding to laminin/merosin. Protein kinase C inhibitors inhibited Mn2+ -enhanced thymocyte binding, suggesting that protein kinase C activity is crucial for the binding. We also showed that both immobilized laminin and merosin have costimulatory function for anti-CD3-induced thymocyte proliferation, and both anti-alpha3 and anti alpha6 mAbs can block this proliferative response. The cooperative function of alpha3 beta1 and alpha6 beta1 evidenced in the laminin/merosin binding and proliferation assays suggests that thymocyte-merosin interactions may play an important role in thymic T cell development. We demonstrated that the anti-mouse integrin beta1 chain mAb KMI6 selectively recognizes a beta1 epitope that is constitutively expressed by certain immature thymocytes and is induced only slightly on mature thymocytes and peripheral T cells by activation with Con A. Because virtually all cells examined expressed beta1 integrins on their surface, expression of the KMI6 epitope is T cell differentiation stage specific. Most CD3-4-8- thymocytes were KMI6+, with the lowest level of staining observed on the earliest CD44+ IL-2R- cells within this subset. Expression was down-regulated during the CD3-4-8- to CD3-4-8+ transition, and lost by the CD4+8+ stage. Mature single positive thymocytes and resting peripheral T cells were also KM16-. The unique selectivity of KMI6 recognition of beta1 integrins, and its selective enhancement of ligand binding suggest that beta1integrin structure and factors that regulate beta1 integrin binding are correlated with the stage of T cell differentiation. The vitronectin receptor (VNR) functions as a co-stimulatory molecule for the activation of a subset of Vgamma1.1/Cgamma4-bearing gamma/delta T cells. We addressed the question of whether stimulation of these T cells requires both engagement of the VNR by ECM proteins and engagement of the TCR by its Ag. We introduced a TCR- but VNR+mutant T cell hybridoma, TG40, a chimeric molecule that contains the cytoplasmic tail of the TCR zeta-chain fused to the cytoplasmic and transmembrane region of either human CD8 or human CD25. The transfectants expressing the chimeric molecules secreted IL-2 constitutively when the VNR was engaged with ECM proteins. This demonstrates that the binding of the VNR to ECM protein in the presence of the zeta-chain is sufficient to induce cytokine secretion by T cells and does not require recognition of an Ag by the TCR. Such integrin-mediated, Ag-independent activation of T cells may play a critical role in the potentiation of inflammatory responses.
