CALPAIN ACTIVATION IN APOPTOSIS

Programmed cell death is an active process wherein the cell initiates a sequence of events culminating in the fragmentation of its DNA, nucl ear collapse, and disintegration of the cell into small, membrane-boun d apoptotic bodies. Examination of the death program in various models has shown common themes, including a rise in cytoplasmic calcium, cyt oskeletal changes, and redistribution of membrane lipids. The calcium- dependent neutral protease calpain has putative roles in cytoskeletal and membrane changes in other cellular processes; this fact led us to test the role of calpain in a well-known model of apoptotic cell death , that of thymocytes after treatment with dexamethasone. Assays for ca lcium-dependent proteolysis in thymocyte extracts reveal a rise in act ivity with a peak at about 1 hr of incubation with dexamethasone, fall ing to background at approximately 2 hr. Western blots indicate autoly tic cleavage of the proenzyme precursor to the calpain I isozyme, prov iding additional evidence for calpain activation. We have also found t hat apoptosis in thymocytes, whether induced by dexamethasone or by lo w-level irradiation, is blocked by specific inhibitors of calpain. Apo ptosis of metamyelocytes incubated with cycloheximide is also blocked by calpain inhibitors. These studies suggest a required role for calpa in in both ''induction'' and ''release'' models of apoptotic cell deat h. (C) 1994 Wiley-Liss, Inc.