EUKARYOTIC TRANSLATION INITIATION-FACTOR 4G IS TARGETED FOR PROTEOLYTIC CLEAVAGE BY CASPASE-3 DURING INHIBITION OF TRANSLATION IN APOPTOTICCELLS

Although much is known about the multiple mechanisms which induce apop tosis, comparatively little is understood concerning the execution pha se of apoptosis and the mechanism(s) of cell killing. Several reports have demonstrated that cellular translation is shut off during apoptos is; however, details of the mechanism of translation inhibition are la cking. Translation initiation factor 4G (eIF4G) is a crucial protein r equired for binding cellular mRNA to ribosomes and is known to be clea ved as the central part of the mechanism of host translation shutoff e xerted by several animal viruses. Treatment of HeLa cells with the apo ptosis inducers cisplatin and etoposide resulted in cleavage of eIF4G, and the extent of its cleavage correlated with the onset and extent o f observed inhibition of cellular translation, The eIF4G-specific clea vage activity could be measured in cell lysates in vitro and was inhib ited by the caspase inhibitor Ac-DEVD-CHO at nanomolar concentrations, A combination of in vivo and in vitro inhibitor studies suggest the i nvolvement of one or more caspases in the activation and execution of eIF4G cleavage. Furthermore recombinant human caspase 3 was expressed in bacteria, and when incubated with HeLa cell lysates, was shown to p roduce the same eIF4G cleavage products as those observed in apoptotic cells. In addition, purified caspase 3 caused cleavage of purified eI F4G, demonstrating that eIF4G could serve as a substrate for caspase 3 , Taken together, these data suggest that cellular translation is spec ifically inhibited during apoptosis by a mechanism involving cleavage of eIF4G, an event dependent on caspase activity.