To identify proteins (enzymes) involved in the T-cell receptor (TCR)-triggered mechano-biochemical activities of cytotoxic T-lymphocytes (CTL) we studied the biochemical changes in cloned and polyclonal CTL after occupation of functionally important surface proteins by antigens or mAb. The ultimate goal of these studies is to understand molecular mechanisms of cytotoxicity and to design specific peptidic reagents that will modulate the intensity of effector functions of lymphocytes. Biochemistry and Immunopharmacology - Since in our earlier studies we implicated Ca2+/Calmodulin dependent, cyclic nucleotide dependent protein kinases, protein kinase C into """"""""ON"""""""" and """"""""OFF' signaling in CTL, we were designing and testing different synthetic peptides substrates, pseudosubstrates and inhibitors of these enzymes to affect effector functions of lymphocytes. We demonstrated the feasibility of such approach in modulation of the functional responses of intact, unpermeabilized cells. Molecular Mechanisms of Cytotoxicity. We proposed the new model of cell mediated cytotoxicity, the """"""""Extracellular ATP"""""""" hypothesis and demonstrated that ATP destroys tumor target cells while CTL themselves are protected by the high levels of ecto-ATPase activity. ATP is accumulated by CTL in response to the TCR crosslinking and the addition of ATP-degrading enzymes blocks cytotoxicity. In control experiments removal of ATP with enzyme does not interfere with the activation of CTL by targets (granule exocytosis response). The presence of ecto-protein kinase activities on both CTL and target cells is demonstrated.
|Kojima, H; Toda, M; Sitkovsky, M V (2000) Comparison of Fas- versus perforin-mediated pathways of cytotoxicity in TCR- and Thy-1-activated murine T cells. Int Immunol 12:365-74|