DEVELOPMENTAL CONTROL OF T-CELL RECEPTOR INTERNALIZATION

Since TCR/CD3 modulation may be involved in induction of T cell tolera nce to self antigens, we compared ligand-induced TCR/CD3 internalizati on by a CTL clone and by immature thymocytes and mature T cells from m ice bearing the same TCR alpha beta as transgene. The ligand used is a monoclonal antibody (mAb) specific for the receptor expressed by the clone and transgenic mice (anti-Ti mAb). CD8(+) splenocytes triggered by anti-Ti mAb internalize the ligand-TCR/CD3 complex at a low rate, t hrough a mechanism inhibited by the protein tyrosine kinase (PTK) inhi bitor genistein and by staurosporine, a potent but non selective prote in kinase C (PC) inhibitor. This pattern of inhibition was similar to that observed in the CTL clone. Anti-Ti mAb induced TCR/CD3 internaliz ation in CD4(+)CD8(+) thymocytes at a high rate, through a mechanism w hich was insensitive to either genistein or staurosporine. In the CTL clone, genistein was shown to inhibit TCR/CD3 surface redistribution p receeding internalization. To characterize the PTK possibly involved i n this step, we analyzed TCR/CD3 associated kinases in mature T spleno cytes and thymocytes. Kinase activities present in anti-Ti mAb immunop recipitates phosphorylated the CD3 components gamma, delta, epsilon, a nd zeta in both cell types although the intensity was stronger in sple nic than in thymocyte extracts, whereas the phosphorylation of 70, 14 and 12kD substrates was more pronounced in thymocytes than in splenocy tes. Comparable amounts of CD3 components were coprecipitated with and phosphorylated by p56lck and p59fyn respectively, in both cell types.