Fang Han1,2, Shuang Liu1,3, Jingjing Jing1, Hao Li1, Yuan Yuan1, Li-Ping Sun1. 1. Tumor Etiology and Screening Department of Cancer Institute and General Surgery, The First Hospital of China Medical University, and Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department, Shenyang 110001, People's Republic of China. 2. Hepatobiliary and Pancreatic Surgery, Minimal Invasive Surgery, Zhejiang Provincial People's Hospital, Hangzhou Medical College, Hangzhou 310014, People's Republic of China. 3. Department of Oncology, Shanxi Provincial Tumor Hospital, Xi'an 710076, People's Republic of China.
Abstract
BACKGROUND: Ring 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 METHODS: A 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. RESULTS: The 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 (P M5 = 0.008, P M27 = 0.003, HRM5(-148) = 2.000 (1.201,3.332), HRM27(-26)=2.389 (1.336,4.271)), which could be independent factors of prognosis. CONCLUSION: By 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.
BACKGROUND: Ring 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 METHODS: A 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. RESULTS: The 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 (P M5 = 0.008, P M27 = 0.003, HRM5(-148) = 2.000 (1.201,3.332), HRM27(-26)=2.389 (1.336,4.271)), which could be independent factors of prognosis. CONCLUSION: By 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.
Authors: Anna Bartlett; Ronan C O'Malley; Shao-Shan Carol Huang; Mary Galli; Joseph R Nery; Andrea Gallavotti; Joseph R Ecker Journal: Nat Protoc Date: 2017-07-20 Impact factor: 13.491
Authors: Michael Rose; Sarah Bringezu; Laura Godfrey; David Fiedler; Nadine T Gaisa; Maximilian Koch; Christian Bach; Susanne Füssel; Alexander Herr; Doreen Hübner; Jörg Ellinger; David Pfister; Ruth Knüchel; Manfred P Wirth; Manja Böhme; Edgar Dahl Journal: Int J Mol Sci Date: 2020-02-07 Impact factor: 5.923