| Research Abstract |
(1) DNA fragments aberrantly methylated in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were searched for using a genome-wide screening technique, methylation-sensitive-representational difference analysis (MS-RDA). However, no silenced genes were identified. (2) Using surgical specimens of human gastric cancers, we identified methylation of a region within the INSIGI promoter CpG island and decreased INSIGI expression in association with the methylation. We also identified methylation of a CpG island covering exon 8 of p4lArc, a subunit of the Arp2/3 complex. Although its methylation was not associated with its reduced expression, expression of seven subunits of the Arp2/3 complex was frequently decreased in gastric cancers. (3) MS-RDA was performed using human gastric cancer cell lines, and we identified nine CpG islands that were in 5' regions and densely methylated. Treatment of gastric cancer cell lines with a demethylating agent, 5-aza-2'-deoxycitidine, confirmed that their downstream genes, LOX, HRASLS, FLNc, HAND1, Thrombomodulin, bA305P22.2.3, a homologue of RIKEN2210016F16, FL 132130, and PGAR, were silenced by the methylation of the 5 CpG islands. The first five genes were methylated in 12-17 (29-41 %) of 41 surgical specimens, and the methylation was associated with reduced gene expression. (4) LOX and HRASLS, which were known to induce reversion of ras-transformants, were introduced into gastric cancer cell lines. LOX was found to suppress cell growth, the number of colonies in soft agar, and sizes of tumors transplanted into nude mice, while HRASLS did not. (5) Frequent methylation of CpG islands and frequent demethylation of normally methylated CpG islands were analyzed in 93 gastric cancer surgical specimens. Frequent methylation and frequent demethylation were found in different sets of gastric cancers.
|
