Down-regulation of connexin 32 gene expression through DNA methylation in a human renal cell carcinoma cell.
BACKGROUND
We have recently reported that connexin (Cx) 32 is down-regulated in a human renal cell carcinoma (RCC) cell (Caki-2 cell).


OBJECTIVE
We postulated that the down-regulation of Cx32 gene in the RCC cell is due to hypermethylation of its promoter region.


METHODS
We estimated methylation status in the promoter region of Cx32 gene in the RCC cell by DNA digestion with methylation-sensitive restriction enzyme and PCR, and methylation-specific PCR (MSP). We also checked the recovery of Cx32 gene expression in the RCC cell treated with a DNA methyltranferase inhibitor, 5-Aza-2'-deoxycytidine (5-Aza-CdR).


RESULTS
Treatment with 5-Aza-CdR resulted in induction of Cx32 expression in the RCC cell. Hypermethylation of the Cx32 promoter region in the RCC cell was confirmed by DNA digestion with methylation-sensitive restriction enzyme and PCR, and MSP.


CONCLUSIONS
We suggest that hypermethylation in the promoter region is a mechanism for the Cx32 gene repression in the RCC cell.
Hirai
Ayako
A
Department of Veterinary Pharmacology, College of Bioresource Sciences, Nihon University, Kanagawa, Japan.
Yano
Tomohiro
T
Nishikawa
Kazuhiro
K
Suzuki
Kazuyuki
K
Asano
Ryuji
R
Satoh
Haruna
H
Hagiwara
Kiyokazu
K
Yamasaki
Hiroshi
H
D000704:Analysis of Variance; D001374:Azacitidine / Q000494:pharmacology; D002292:Carcinoma, Renal Cell / Q000235:genetics*; D002450:Cell Communication; D017630:Connexins / Q000235:genetics*; D019175:DNA Methylation; D015536:Down-Regulation; D015870:Gene Expression; D006801:Humans; D015151:Immunoblotting; D007680:Kidney Neoplasms / Q000235:genetics*; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D000097002:Gap Junction beta-1 Protein
D001374:Azacitidine; D017630:Connexins; D000097002:Gap Junction beta-1 Protein