NUCLEAR EVENTS OF APOPTOSIS IN-VITRO IN CELL-FREE MITOTIC EXTRACTS - A MODEL SYSTEM FOR ANALYSIS OF THE ACTIVE PHASE OF APOPTOSIS

We have developed a cell-free system that induces the morphological tr ansformations characteristic of apoptosis in isolated nuclei. The syst em uses extracts prepared from mitotic chicken hepatoma cells followin g a sequential S phase/M phase synchronization. When nuclei are added to these extracts, the chromatin becomes highly condensed into spheric al domains that ultimately extrude through the nuclear envelope, formi ng apoptotic bodies. The process is highly synchronous, and the struct ural changes are completed within 60 min. Coincident with these morpho logical changes, the nuclear DNA is cleaved into a nucleosomal ladder. Both processes are inhibited by Zn2+, an inhibitor of apoptosis in in tact cells. Nuclear lamina disassembly accompanies these structural ch anges in added nuclei, and we show that lamina disassembly is a charac teristic feature of apoptosis in intact cells of mouse, human and chic ken. This system may provide a powerful means of dissecting the bioche mical mechanisms underlying the final stages of apoptosis.