Up-regulation of connexin 32 gene by 5-aza-2'-deoxycytidine enhances vinblastine-induced cytotoxicity in human renal carcinoma cells via the activation of JNK signalling.
Enforced expression of connexin (Cx) 32 gene, a member of gap junction gene family and a tumor suppressor gene in human renal cell carcinoma (RCC), enhanced vinblastine (VBL)-induced cytotoxicity on RCC cells, due to the suppression of multidrug resistance 1 (MDR1) gene product, P-glycoprotein (P-gp). Also, Cx32 gene in RCC is silenced by hypermethylation of CpG islands in a promoter region of the Cx gene. In this study, we investigated if a DNA demethylating agent, 5-aza-2'-deoxycytidine (5-Aza) could enhance susceptibility of RCC cells (Caki-1) to VBL. We found that 5-Aza treatment up-regulated Cx32 in Caki-1 cells, and the induction of the Cx led to the suppression of P-gp through inhibition of Src and subsequent activation of c-Jun NH(2)-terminal kinase (JNK). Moreover, increased transcription activity of c-Jun by the JNK activation contributed to the down-regulation of MDR1, thus indicating a central role of JNK signalling to suppress P-gp level in 5-Aza-treated Caki-1 cells. Chemical sensitivity to VBL in Caki-1 cells was increased by 5-Aza pre-treatment, and this effect was abrogated by short interfering RNA (siRNA)-mediated knockdown of Cx32. Furthermore, co-treatment of 5-Aza or a P-gp inhibitor with VBL drastically enhanced JNK activation comparing to only VBL treatment in Caki-1 cells. These results suggest that the restoration of Cx32 by 5-Aza pre-treatment improves chemical tolerance on VBL in Caki-1 cells and that the JNK activation is a key factor to induce the effect.
National Institute of Health and Nutrition, Tokyo, Japan. ya.tomohiro@gmail.com
Y
Y
Takano
H
H
Iwata
Y
Y
Yano
M
M
Miyazawa
N
N
Virgona
H
H
Sato
K
K
Ueno
T
T
Yano
D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002292:Carcinoma, Renal Cell / Q000473:pathology*; D049109:Cell Proliferation / Q000187:drug effects; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D000077209:Decitabine; D003841:Deoxycytidine / Q000494:pharmacology; D015536:Down-Regulation; D004789:Enzyme Activation; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D007680:Kidney Neoplasms / Q000378:metabolism / Q000473:pathology*; D015398:Signal Transduction / Q000187:drug effects* / Q000528:radiation effects; D015533:Transcriptional Activation / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D014747:Vinblastine / Q000494:pharmacology*; D000097002:Gap Junction beta-1 Protein
D001374:Azacitidine; D017630:Connexins; D000077209:Decitabine; D003841:Deoxycytidine; D048031:JNK Mitogen-Activated Protein Kinases; D014747:Vinblastine; D000097002:Gap Junction beta-1 Protein