Inhibition of the JNK signalling pathway enhances proteasome inhibitor-induced apoptosis of kidney cancer cells by suppression of BAG3 expression.
OBJECTIVE
Proteasome inhibitors represent a novel class of anti-tumour agents that have clinical efficacy against haematological and solid cancers. The anti-apoptotic protein BAG3 is a member of the Bcl-2-associated athanogene family. We have previously shown that BAG3 is up-regulated after exposure to proteasome inhibitors and that inhibition of BAG3 sensitized cells to apoptosis induced by proteasome inhibition. However, the mechanisms by which proteasome inhibition induced BAG3 expression remained unclear and the present experiments were designed to elucidate these mechanisms.


METHODS
Effects of the proteasome inhibitor MG132 on activation of mitogenic signalling pathways were evaluated in kidney cancer cells (A498, Caki1, Caki2), with Western blotting. Specific inhibitors against individual mitogenic signalling pathways, real-time reverse transcription-polymerase chain reaction and luciferase reporter assays were used to investigate the roles of mitogenic signalling pathways in BAG3 induction after proteasome inhibition. Cell death was evaluated using Annexin V/propidium iodide staining and subsequent FACS.


RESULTS
MG132 activated several key mitogenic signalling pathways including extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) activities. Induction of BAG3 by MG132 was inhibited by blocking JNK, but not ERK1/2 and p38 MAPK signalling pathways. In addition, SP600125 and dominant-negative JNK1 suppressed BAG3 promoter-driven reporter gene expression. Furthermore, activation of the JNK pathway induced BAG in kidney cancer cells after treatment with MG132.


CONCLUSIONS
Our results suggested that the JNK pathway was associated with the protective response against proteasome inhibition, by mediating induction of BAG3.
Wang
Hua-Qin
HQ
Department of Biochemistry & Molecular Biology, China Medical University, Shenyang, China. wanghq_doctor@hotmail.com
Liu
Bao-Qin
BQ
Gao
Yan-Yan
YY
Meng
Xin
X
Guan
Yifu
Y
Zhang
Hai-Yan
HY
Du
Zhen-Xian
ZX
D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis*; D000873:Anthracenes / Q000494:pharmacology*; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D045744:Cell Line, Tumor; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases / Q000502:physiology*; D007680:Kidney Neoplasms; D007976:Leupeptins / Q000494:pharmacology; D048055:Mitogen-Activated Protein Kinase 8 / Q000235:genetics; D061988:Proteasome Inhibitors; D011725:Pyridines / Q000494:pharmacology; D015398:Signal Transduction; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors
D048868:Adaptor Proteins, Signal Transducing; D000873:Anthracenes; D051017:Apoptosis Regulatory Proteins; C414431:BAG3 protein, human; D005419:Flavonoids; D007093:Imidazoles; D007976:Leupeptins; D061988:Proteasome Inhibitors; D011725:Pyridines; C432165:pyrazolanthrone; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D048055:Mitogen-Activated Protein Kinase 8; D048051:p38 Mitogen-Activated Protein Kinases; C093642:SB 203580; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one
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