Correlation of calcineurin phosphatase activity and programmed cell death in murine T cell hybridomas.
Ligation of T cell receptor/CD3 complexes induces programmed cell death, or apoptosis, in immature thymocytes and many T cell hybridomas. While it has been demonstrated that T cell receptor-mediated apoptosis requires an increase in intracellular calcium concentration, the specific calcium-dependent signalling events leading to cell death are poorly defined. We have previously shown that T cell receptor/CD3-mediated induction of apoptosis in a murine T cell hybridoma is inhibited by the immunosuppressive drugs cyclosporin A (CsA) and FK506. Recently, it has been determined that these agents inhibit the activity of calcineurin, a calcium- and calmodulin-dependent serine/threonine phosphatase. Using an assay which measures calcineurin activity in cell lysates, we find that calcineurin-dependent dephosphorylation of a phosphopeptide substrate is potently inhibited in hybridomas treated with CsA or FK506. Drug dose-response analyses indicate that the level of cellular calcineurin activity correlates closely with the ability of these cells to undergo apoptosis. Thus, calcineurin appears to be a critical mediator of T cell receptor/CD3 signalling leading to programmed cell death in T cell hybridomas.
Fruman
D A
DA
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA 02115.
Mather
P E
PE
Burakoff
S J
SJ
Bierer
B E
BE
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D017252:CD3 Complex / Q000502:physiology; D019703:Calcineurin; D002118:Calcium / Q000502:physiology; D002148:Calmodulin-Binding Proteins / Q000032:analysis*; D016572:Cyclosporine / Q000494:pharmacology; D006801:Humans; D006825:Hybridomas / Q000201:enzymology / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000032:analysis*; D011090:Polyenes / Q000494:pharmacology; D020123:Sirolimus; D013601:T-Lymphocytes / Q000201:enzymology / Q000502:physiology*; D016559:Tacrolimus / Q000494:pharmacology
D017252:CD3 Complex; D002148:Calmodulin-Binding Proteins; D011090:Polyenes; D016572:Cyclosporine; D019703:Calcineurin; D010749:Phosphoprotein Phosphatases; D002118:Calcium; D020123:Sirolimus; D016559:Tacrolimus