Zhong-Ming Huang1, Hai Wang1, Zhi-Gang Ji2. 1. Department of Urology, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, No.1 Shuaifuyuan Wangfujing, Dongcheng, Beijing, 100730, China. 2. Department of Urology, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, No.1 Shuaifuyuan Wangfujing, Dongcheng, Beijing, 100730, China. jizhigang@pumch.cn.
Abstract
OBJECTIVE: The malignant transformation of normal bladder cells (SV-HUC-1) was induced by arsenite to explore the possible mechanism of circRNA-100284 influencing bladder cancer cell proliferation. METHODS: Normal bladder SV-HUC-1 cells were cultured with 2 μM arsenite to induce malignant transformation. After 0, 3, 6, 12, and 24 h of culture, the expression level of circRNA-100284 in cells was detected by quantitative real-time PCR. Western blotting assays were used to detect the expression levels of EZH2 and cyclin-D1 proteins in cells treated with different media. Cell cycle was analyzed by flow cytometry. In addition, through cell transfection and CCK-8 experiments, the effect and mechanism of circRNA-100284 targeting microRNA-217 on proliferation was determined. The interaction between HSP70 methylation and Aurora-B was determined by Western blotting and immunoprecipitation experiments. RESULTS: With prolonged contact time with arsenite, the expression level of circRNA-100284 in cells increased continuously (P < 0.05). Western blotting assays showed that the expression levels of EZH2 and cyclin-D1 proteins in arsenite-transformed cells increased. Flow cytometry and CCK-8 showed that circRNA-100284 accelerated cell cycle transition and cell proliferation through miR-217. Finally, after culturing human bladder cancer T24 cells, combined with immunoprecipitation and in vitro kinase experiments, it was found that K561- dimethyl HSP70 activated Aurora-B, thus promoting the proliferation of bladder cancer cells. CONCLUSION: CircRNA-100284 activates aurora kinase B by inducing methylation of HSP70 via microRNA-217 to promote the proliferation of bladder cancer cells.
OBJECTIVE: The malignant transformation of normal bladder cells (SV-HUC-1) was induced by arsenite to explore the possible mechanism of circRNA-100284 influencing bladder cancer cell proliferation. METHODS: Normal bladder SV-HUC-1 cells were cultured with 2 μM arsenite to induce malignant transformation. After 0, 3, 6, 12, and 24 h of culture, the expression level of circRNA-100284 in cells was detected by quantitative real-time PCR. Western blotting assays were used to detect the expression levels of EZH2 and cyclin-D1 proteins in cells treated with different media. Cell cycle was analyzed by flow cytometry. In addition, through cell transfection and CCK-8 experiments, the effect and mechanism of circRNA-100284 targeting microRNA-217 on proliferation was determined. The interaction between HSP70 methylation and Aurora-B was determined by Western blotting and immunoprecipitation experiments. RESULTS: With prolonged contact time with arsenite, the expression level of circRNA-100284 in cells increased continuously (P < 0.05). Western blotting assays showed that the expression levels of EZH2 and cyclin-D1 proteins in arsenite-transformed cells increased. Flow cytometry and CCK-8 showed that circRNA-100284 accelerated cell cycle transition and cell proliferation through miR-217. Finally, after culturing humanbladder cancer T24 cells, combined with immunoprecipitation and in vitro kinase experiments, it was found that K561- dimethyl HSP70 activated Aurora-B, thus promoting the proliferation of bladder cancer cells. CONCLUSION: CircRNA-100284 activates aurora kinase B by inducing methylation of HSP70 via microRNA-217 to promote the proliferation of bladder cancer cells.
Authors: T Adhikary; D T Brandt; K Kaddatz; J Stockert; S Naruhn; W Meissner; F Finkernagel; J Obert; S Lieber; M Scharfe; M Jarek; P M Toth; F Scheer; W E Diederich; S Reinartz; R Grosse; S Müller-Brüsselbach; R Müller Journal: Oncogene Date: 2012-12-03 Impact factor: 9.867