Characterization of distinct consecutive phases in non-genotoxic p53-induced apoptosis of Ewing tumor cells and the rate-limiting role of caspase 8.

Abstract:

:To dissect the p53-dependent apoptotic pathway, events following induction of temperature sensitive (ts) p53val138 were studied in a Ewing tumor cell line. Transcriptional deregulation of p53 targets first observable after 1 h at 32 degrees C preceded activation of caspases and the break-down of mitochondrial respiratory activity. Activation of caspases was first observed 4 h after p53 induction. Using peptide inhibitors we identified activation of caspase 8 upstream of caspases-9 and -3. Although the caspase 8 specific inhibitor z-IETD.fmk did not affect translocation of BAX to the mitochondrial membrane and cytochrome C release it almost completely blocked cleavage of the prototype caspase substrate PARP and DNA fragmentation while enforcing mitochondrial depolarization and production of reactive oxygene species (ROS). Activation of caspase 8 did not involve death-domain receptor signaling. Expression of BCL2 only partially suppressed caspase activation but blocked apoptosis. Replacement of the N-terminus of p53val138 by the related VP16 transactivation domain created a ts p53 with a tanscriptional activity indistinguishable from p53val138 until the time of caspase activation. However, the VP16 - p53 fusion failed to trigger caspases and subsequent induction of the ROS producing gene pig3 paralleled by complete loss of apoptotic activity. These results indicate that p53-dependent transcriptional deregulation, triggering of the caspase cascade and the mitochondrial break-down occur in a timely ordered sequence coordinated by the genuine p53 amino terminus and suggest caspase 8 and PIG3 as key regulatory elements in this process. Oncogene (2000) 19, 4096 - 4107

journal_name

Oncogene

journal_title

Oncogene

authors

Kovar H,Jug G,Printz D,Bartl S,Schmid G,Wesierska-Gadek J

doi

10.1038/sj.onc.1203780

subject

Has Abstract

pub_date

2000-08-24 00:00:00

pages

4096-107

issue

36

eissn

0950-9232

issn

1476-5594

journal_volume

19

pub_type

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