MICROINJECTED CATALYTIC SUBUNIT OF CAMP-DEPENDENT PROTEIN-KINASE INDUCES APOPTOSIS IN MYELOID-LEUKEMIA (IPC-81) CELLS

Rat IPC-81 promyelocytic leukemia cells responded to cAMP analog by undergoing apoptotic cell death both when anchored to fibronectin and when free in the medium. The protein kinase C stimulator 12-O-tetradecanoylphorbol 13-acetate enhanced the anchoring to substratum without impeding cAMP-induced cell death. The immobilized cells could be microinjected. This made it possible to study the effect on apoptosis of microinjected catalytic (Cα) and regulatory (RIαD199) subunits of cAMP-dependent protein kinase as well as of phosphatase inhibitors. Microinjection of Cα reproduced the morphological effects of cAMP, including nuclear fragmentation. RIαD199 blocked the effect of Cα. Injection of microcystin-LR, which inhibits protein phosphatases 1 and 2A, led to pronounced apoptoid changes of the leukemia cells, but failed to produce nuclear fragmentation. Microinjection of peptide inhibitors (“inhibitor 1” and “inhibitor 2”) specific for phosphatase 1 had no effect on cell morphology. The failure of the phosphatase inhibitors to reproduce completely the effect of the C subunit underscores the specificity of action of the latter. © 1993 Academic Press, Inc.