Tunicamycin sensitizes human melanoma cells to tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis by up-regulation of TRAIL-R2 via the unfolded protein response.
We have reported previously low expression of death receptors for tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) in fresh isolates and tissue sections of melanoma. This seemed to correlate with relative resistance of freshly isolated melanoma cells to TRAIL-induced apoptosis. We show in this study that the endoplasmic reticulum (ER) stress inducer, tunicamycin, selectively up-regulated the cell surface expression of TRAIL-R2, but not other members of the TNF receptor family, and enhanced TRAIL-induced apoptosis in cultured melanoma cells and fresh melanoma isolates. Tunicamycin-mediated sensitization of melanoma cells to TRAIL-induced apoptosis was associated with increased activation of the caspase cascade and reduction in mitochondrial membrane potential and was inhibited by a recombinant TRAIL-R2/Fc chimeric protein. Up-regulation of TRAIL-R2 on the melanoma cell surface was associated with increased transcription of TRAIL-R2 and its total protein levels. Two signaling pathways of the ER stress-induced unfolded protein response mediated by inositol-requiring transmembrane kinase and endonuclease 1alpha (IRE1alpha) and activation of transcription factor 6 (ATF6), respectively, seemed to be involved. In one melanoma line, there was clear evidence of activation of the IRE1alpha pathway, and small interfering RNA (siRNA) knockdown of IRE1alpha substantially reduced the up-regulation of TRAIL-R2. Similarly, there was evidence for the activation of the ATF6 pathway, and siRNA knockdown of ATF6 had a delayed effect on TRAIL-R2 expression in one but not another melanoma cell line. Moreover, the transcription factor CCAAT/enhancer-binding protein homologous protein seemed to be involved in the up-regulation of TRAIL-R2 by tunicamycin, but its role varied between different melanoma lines. Taken together, our results suggest that agents that induce ER stress may enhance TRAIL-R2 expression and increase the therapeutic response to TRAIL in melanoma.
Jiang
Chen Chen
CC
Immunology and Oncology Unit, Newcastle Misericordiae Hospital, Newcastle, NSW, Australia.
Chen
Li Hua
LH
Gillespie
Susan
S
Kiejda
Kelly A
KA
Mhaidat
Nizar
N
Wang
Yu Fang
YF
Thorne
Rick
R
Zhang
Xu Dong
XD
Hersey
Peter
P
D051702:Activating Transcription Factor 6 / Q000187:drug effects / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D020869:Gene Expression Profiling; D006801:Humans; D008545:Melanoma / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D018613:Microscopy, Confocal; D051336:Mitochondrial Membranes / Q000187:drug effects; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D014157:Transcription Factors / Q000187:drug effects / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology*; D015854:Up-Regulation
C497111:ATF6 protein, human; D051702:Activating Transcription Factor 6; D000900:Anti-Bacterial Agents; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D053221:TNF-Related Apoptosis-Inducing Ligand; D014157:Transcription Factors; D014415:Tunicamycin