Degradation of retinoid X receptor alpha by TPA through proteasome pathway in gastric cancer cells.
OBJECTIVE
To investigate and determine the mechanism and signal pathway of tetradecanoylphorbol-1, 3-acetate (TPA) in degradation of RXRalpha.


METHODS
Gastric cancer cell line, BGC-823 was used in the experiments. The expression level of RXRalpha protein was detected by Western blot. Nuclear and cytoplasmic protein fractions were prepared through lysis of cell and centrifugation. Localization and translocation of RXRalpha were observed under laser-scanning confocal microscope through labeling specific anti-RXRalpha antibody and corresponding immunofluorescent antibody as secondary antibody. Different inhibitors were used as required.


RESULTS
In BGC-823 cells, RXRalpha was expressed in the nucleus. When cells were treated with TPA, expression of RXRalpha was repressed in a time-dependent and TPA-concentration-dependent manner. Meanwhile, translocation of RXRalpha from the nucleus to the cytoplasm occurred, also in a time-dependent manner. When cells were pre-incubated with proteasome inhibitor MG132 for 3 hrs, followed by TPA for another 12 hrs, TPA-induced RXRalpha degradation was inhibited. Further observation of RXRalpha translocation in the presence of MG132 showed that MG-132 could block TPA-induced RXRalpha redistribution. Conversely, when RXRalpha translocation was inhibited by LMB, an inhibitor for blocking protein export from the nucleus, TPA could not repress expression of RXRalpha.


CONCLUSIONS
TPA could induce the degradation of RXRalpha protein in BGC-823 cells, and this degradation is time- and TPA-concentration-dependent. Furthermore, the degradation of RXRalpha by TPA is via a proteasome pathway and associated with RXRalphatranslocation from the nucleus to the cytoplasm.
Ye
Xiao-Feng
XF
Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Xiamen 361005, Fujian Province, China.
Liu
Su
S
Wu
Qiao
Q
Lin
Xiao-Feng
XF
Zhang
Bing
B
Wu
Jia-Fa
JF
Zhang
Ming-Qing
MQ
Su
Wen-Jin
WJ
D002273:Carcinogens / Q000494:pharmacology*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D006801:Humans; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D018168:Receptors, Retinoic Acid / Q000187:drug effects* / Q000378:metabolism*; D047488:Retinoid X Receptors; D013274:Stomach Neoplasms / Q000378:metabolism* / Q000473:pathology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014157:Transcription Factors / Q000187:drug effects* / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D002273:Carcinogens; D009097:Multienzyme Complexes; D018168:Receptors, Retinoic Acid; D047488:Retinoid X Receptors; D014157:Transcription Factors; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D013755:Tetradecanoylphorbol Acetate