Abstract
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an inducer of cancer cell death that holds promise in cancer therapy. Cancer cells are more susceptible than normal cells to the cell-death-inducing effects of TRAIL. However, a variety of cancer cells are resistant to TRAIL through complex mechanisms. Here, we investigate the effects of inhibition of eukaryotic initiation factor 2 subunit α (eIF2α) dephosphorylation on TRAIL-induced apoptosis in hepatoma cells. Treatment of hepatoma cells with salubrinal, an inhibitor of eIF2α dephosphorylation, enhances TRAIL-induced eIF2α phosphorylation, CCAAT/enhancer-binding protein homologous protein (CHOP) expression and caspase activation. Salubrinal enhances TRAIL-induced apoptosis, which could be abrogated by caspase inhibitor. Overexpression of phosphomimetic eIF2α (S51D) enhances TRAIL-induced CHOP expression, caspase 7 and PARP cleavage and apoptosis. By contrast, overexpression of phosphodeficient eIF2α (S51A) abrogates the stimulation of TRAIL-induced apoptosis by salubrinal. Moreover, knockdown of growth arrest and DNA damage-inducible protein 34 (GADD34), which recruits protein phosphatase 1 to dephosphorylate eIF2α, enhances TRAIL-induced eIF2α phosphorylation, CHOP expression, caspase activation and apoptosis. Furthermore, the sensitization of hepatoma cells to TRAIL by salubrinal is dependent on CHOP. Knockdown of CHOP abrogates the stimulation of TRAIL-induced caspase activation and apoptosis by salubrinal. Combination of salubrinal and TRAIL leads to increased expression of Bim, a CHOP-regulated proapoptotic protein. Bim knockdown blunts the stimulatory effect of salubrinal on TRAIL-induced apoptosis. Collectively, these findings suggest that inhibition of eIF2α dephosphorylation may lead to synthetic lethality in TRAIL-treated hepatoma cells.
Highlights
Given that TNF-related apoptosis-inducing ligand (TRAIL) induces Eukaryotic initiation factor 2 subunit a (eIF2a) phosphorylation, we wish to determine the effects of selective inhibition of eIF2a dephosphorylation on TRAIL-induced apoptosis
We investigated whether TRAIL would induce endoplasmic reticulum (ER) stress or eIF2a phosphorylation in hepatoma cells
The results showed that eIF2a phosphorylation was induced by TRAIL in a dose-dependent manner, while GRP78 expression was unaffected by TRAIL (Figures 1b and c)
Summary
TRAILR1 ( known as DR4) and TRAILR2 ( known as DR5). Binding of TRAIL to TRAILR1 or TRAILR2 leads to receptor oligomerization and recruitment of the FAS-associated protein with death domain (FADD) to death domain motifs in the carboxyl terminus of the receptor thereby recruiting caspase-8 or caspase-10 to form a multiprotein death-inducing signaling complex.[2]. Phosphorylation of eIF2a leads to increased synthesis of activating transcription factor 4 (ATF4) thereby increasing the expression of growth arrest and DNA damage-inducible protein 34 (GADD34), which recruits protein phosphatase 1 to eIF2a and dephosphorylates eIF2a.19. This in turn results in the recovery of protein synthesis and restoration of homeostasis. Prolonged or unresolved ER stress, can induce cell death, which may involve eIF2a phosphorylation and increased ATF4 and CCAAT/enhancer-binding protein homologous protein (CHOP) expression. Treatment with salubrinal leads to an increase in TRAIL-induced eIF2a phosphorylation, CHOP and Bim expression. CHOP or Bim knockdown blunts the stimulation of TRAIL-induced apoptosis by salubrinal
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