Identification of High-Frequency Methylation Sites in RNF180 Promoter Region Affecting Expression and Their Relationship with Prognosis of Gastric Cancer.

backgroundRing finger protein 180 (RNF180) is a tumor suppressor gene regulated by promoter methylation. We previously demonstrated that the RNF180 promoter methylation could be a risk factor for gastric cancer (GC); and eight high-frequency hypermethylated CpG sites were associated with GC. However, it is not clear whether these key sites can affect gene expression and involve in prognosis. The aim of this study was to investigate the effects of above CpG sites on the gene expression and prognosis of GC. PATIENTS AND

methodsA total of 164 GC tissues were enrolled and followed up. Tissue samples were used for DNA and RNA isolation. Methylation status of RNF180 was detected using bisulfite sequencing PCR (BSP). Expression levels of RNF180 were detected using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). JASPAR and PROMO databases were used to predict the transcription factors (TFs) binding to the CpG site.

resultsThe methylation in RNF180 promoter region increased and mRNA expression decreased in GC tissue. Correlation analysis revealed that the average methylation rate (AMR) and four CpG sites methylation rate were negatively related to RNF180 expression, including M3(-165)(Chr5:64165942), M5(-148)(Chr5:64,165,959), M7(-133)(Chr5:64,165,974) and M8(-130)(Chr5:64,165,977). Furthermore, the methylation rate of M5(-148)(Chr5:64,165,959) and M27(-26)(Chr5:64,166,081) above 0.3 indicated poor prognosis (

conclusionBy focusing on the methylation sites in the RNF180 promoter region, we identified two high-frequency methylation sites, M5(-148)(Chr5:64,165,959) and M27(-26)(Chr5:64,166,081), which could affect gene expression and predict the prognosis of GC. In the future, the possible molecular mechanism involved needs to be further studied.