CpG hypermethylation of collagen type I alpha 2 contributes to proliferation and migration activity of human bladder cancer.
In our microarray screening of methylated genes in bladder cancer (BC), the collagen type 1 alpha 2 (COL1A2) gene was the most up-regulated among the 30,144 genes screened. We hypothesize that inactivation of the COL1A2 gene through CpG methylation contributes to proliferation and migration activity of human BC. We subjected a bladder cancer cell line (BOY) and 67 BC specimens and 10 normal bladder epitheliums (NBEs) to conventional or real-time methylation quantitative polymerase chain reaction (PCR) and to real-time reverse transcriptase (RT)-PCR. We also established a stable COL1A2 transfectant for evaluating cell proliferation and migration activity. After 5-aza-dC treatment, the expression levels of COL1A2 mRNA transcript markedly increased in BOY. Our cell proliferation assays consistently demonstrated growth inhibition in the COL1A2 transfectant compared with control and wild-type BOY cells (p<0.0001). Wound healing assays also showed significant wound healing inhibition in the COL1A2 transfectant compared to the counterparts (p=0.0016). We demonstrated by bisulfite DNA sequencing that the promoter hypermethylation of COL1A2 was a frequent event in clinical BCs. The methylation index of COL1A2 was significantly higher in the 67 BCs than in the 10 NBEs (p=0.0011). Conversely, COL1A2 mRNA transcript was significantly lower in the BCs than in the NBEs (p=0.0052). The mechanism of COL1A2 down-regulation in BC is through CpG hypermethylation of the promoter region. COL1A2 gene inactivation through CpG hypermethylation may contribute to proliferation and migration activity of BC.
Mori
Katsuhisa
K
Department of Urology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8520, Japan.
Enokida
Hideki
H
Kagara
Ichiro
I
Kawakami
Kazumori
K
Chiyomaru
Takeshi
T
Tatarano
Shuichi
S
Kawahara
Kazuya
K
Nishiyama
Kenryu
K
Seki
Naohiko
N
Nakagawa
Masayuki
M
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion; D002465:Cell Movement; D049109:Cell Proliferation; D003094:Collagen / Q000235:genetics* / Q000378:metabolism; D024042:Collagen Type I; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D000077209:Decitabine; D015536:Down-Regulation; D005260:Female; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D008297:Male; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic; D011471:Prostatic Neoplasms / Q000235:genetics / Q000473:pathology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D001743:Urinary Bladder / Q000378:metabolism / Q000473:pathology; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000473:pathology*
D014408:Biomarkers, Tumor; D024042:Collagen Type I; D012333:RNA, Messenger; D000077209:Decitabine; D003094:Collagen; D001374:Azacitidine