A novel form of DAP5 protein accumulates in apoptotic cells as a result ofcaspase cleavage and internal ribosome entry site-mediated translation

Death-associated protein 5 (DAP5) (also named p97 and NAT1) is a member of the translation initiation factor 4G (eIF4G) family that lacks the eIF4E bi nding site. It was previously implicated in apoptosis, based on the finding that a dominant negative fragment of the protein protected against cell de ath. Here we address its function and two distinct levels of regulation dur ing apoptosis that affect the protein both at translational and posttransla tional levels. DAP5 protein was found to be cleaved at a single caspase cle avage site at position 790, in response to activated Fas or p53, yielding a C-terminal truncated protein of 86 kDa that is capable of generating compl exes with eIF4A and eIF3. Interestingly, while the overall translation rate in apoptotic cells was reduced by 60 to 70%, in accordance with the simult aneous degradation of the two major mediators of cap-dependent translation, eIF4GI and eIF4GII, the translation rate of DAP5 protein was selectively m aintained. An internal ribosome entry site (IRES) element capable of direct ing the translation of a reporter gene when subcloned into a bicistronic ve ctor was identified in the 5' untranslated region of DAP5 mRNA. While cap-d ependent translation from this transfected vector was reduced during Fas-in duced apoptosis, the translation via the DAP5 IRES was selectively maintain ed. Addition of recombinant DAP5/p97 or DAP5/p86 to cell-free systems enhan ced preferentially the translation through the DAP5 IRES, whereas neutraliz ation of the endogenous DAP5 in reticulocyte lysates by adding a dominant n egative DAP5 fragment interfered with this translation. The DAP5/p86 apopto tic form was more potent than DAP5/p97 in these functional assays. Altogeth er, the data suggest that DAP5 is a caspase-activated translation factor wh ich mediates cap-independent translation at least from its own IRES, thus g enerating a positive feedback loop responsible for the continuous translati on of DAP5 during apoptosis.