Essential role of caspase-8 in p53/p73-dependent apoptosis induced by etoposide in head and neck carcinoma cells

Juan Liu,Hiroshi Uematsu,Masa-Aki Ikeda,Nobuo Tsuchida

doi:10.1186/1476-4598-10-95

Abstract

BackgroundCaspase-8 is a key upstream mediator in death receptor-mediated apoptosis and also participates in mitochondria-mediated apoptosis via cleavage of proapoptotic Bid. However, the role of caspase-8 in p53- and p73-dependent apoptosis induced by genotoxic drugs remains unclear. We recently reported that the reconstitution of procaspase-8 is sufficient for sensitizing cisplatin- but not etoposide-induced apoptosis, in chemoresistant and caspase-8 deficient HOC313 head and neck squamous cell carcinoma (HNSCC) cells.ResultsWe show that p53/p73-dependent caspase-8 activation is required for sensitizing etoposide-induced apoptosis by utilizing HOC313 cells carrying a temperature-sensitive p53G285K mutant. Restoration of wild-type p53 function under the permissive conditions, together with etoposide treatment, led to substantial transcriptional activation of proapoptotic Noxa and PUMA, but failed to induce apoptosis. In addition to p53 restoration, caspase-8 reconstitution was needed for sensitization to etoposide-induced apoptosis, mitochondria depolarization, and cleavage of the procaspases-3, and -9. In etoposide-sensitive Ca9-22 cells carrying a temperature-insensitive mutant p53, siRNA-based p73 knockdown blocked etoposide-induced apoptosis and procaspase-8 cleavage. However, induction of p73 protein and up-regulation of Noxa and PUMA, although observed in Ca9-22 cells, were hardly detected in etoposide-treated HOC313 cells under non-permissive conditions, suggesting a contribution of p73 reduction to etoposide resistance in HOC313 cells. Finally, the caspase-9 inhibitor Ac-LEHD-CHO or caspase-9 siRNA blocked etoposide-induced caspase-8 activation, Bid cleavage, and apoptosis in both cell lines, indicating that p53/p73-dependent caspase-8 activation lies downstream of mitochondria.Conclusionswe conclude that p53 and p73 can act as upstream regulators of caspase-8, and that caspase-8 is an essential mediator of the p53/p73-dependent apoptosis induced by etoposide in HNSCC cells. Our data suggest the importance of caspase-8-mediated positive feedback amplification in the p53/p73-dependent apoptosis induced by etoposide in HNSCC cells.

Highlights

Caspase-8 is a key upstream mediator in death receptor-mediated apoptosis and participates in mitochondria-mediated apoptosis via cleavage of proapoptotic Bid
The restoration of wild-type p53 function induced p53target gene expression in etoposide-treated HOC313 cells We previously reported that stable caspase-8 reconstitution sensitizes drug-resistant and caspase-8-deficient HOC313 cells to cisplatin-induced apoptosis [42]
We provide evidence that caspase-8 is an essential mediator of the p53/p73-dependent apoptosis induced by etoposide in head and neck squamous cell carcinoma (HNSCC) cells

Summary

Introduction

Caspase-8 is a key upstream mediator in death receptor-mediated apoptosis and participates in mitochondria-mediated apoptosis via cleavage of proapoptotic Bid. Caspase-8 is predominantly activated by signals from the death receptor pathway, while caspase-9 activation is dependent primarily on the mitochondrial pathway. In both apoptotic pathways, these initiator caspases activate downstream effector caspases (e.g. caspases-3, -6, -7) by a proteolytic cascade, resulting in the cleavage of a variety of cellular substrates involved in apoptosis. Caspase-8 can amplify the death signal by activating the mitochondrial pathway through the cleavage of the BH3-only protein Bid [14,24,25,26,27,28,29,30]. Cleaved Bid (tBid) translocates to the mitochondria and triggers mitochondrial depolarization, leading to cytochrome c release and subsequent caspase-9 activation, by which the activation of caspase-8 initiates a positive feedback loop that amplifies the mitochondrial pathway

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