title: miR-200c inhibits invasion, migration and proliferation of bladder cancer cells through down-regulation of BMI-1 and E2F3.
abstract: BACKGROUND
  MicroRNA-200c (miR-200c) is one of the short noncoding RNAs that play crucial roles in tumorigenesis and tumor progression. It also acts as considerable modulator in the process of epithelial-to-mesenchymal transition (EMT), a cell development regulating process that affects tumor development and metastasis. However, the role of miR-200c in bladder cancer cells and its mechanism has not been well studied. The purpose of this study was to determine the potential role of miR-200c in regulating EMT and how it contributed to bladder cancer cells in invasion, migration and proliferation.
  
  
  METHODS
  Real-time reverse transcription-PCR was used to identify and validate the differential expression of MiR-200c involved in EMT in 4 bladder cancer cell lines and clinical specimens. A list of potential miR-200 direct targets was identified through the TargetScan database. The precursor of miR-200c was over-expressed in UMUC-3 and T24 cells using a lentivirus construct, respectively. Protein expression and signaling pathway modulation were validated through Western blot analysis and confocal microscopy, whereas BMI-1 and E2F3, direct target of miR-200c, were validated by using the wild-type and mutant 3'-untranslated region BMI-1/E2F3 luciferase reporters.
  
  
  RESULTS
  We demonstrate that MiR-200c is down-regulated in bladder cancer specimens compared with adjacent ones in the same patient. Luciferase assays showed that the direct down-regulation of BMI-1 and E2F3 were miR-200c-dependent because mutations in the two putative miR-200c-binding sites have rescued the inhibitory effect. Over-expression of miR-200c in bladder cancer cells resulted in significantly decreased the capacities of cell invasion, migration and proliferation. miR-200c over-expression resulted in conspicuous down-regulation of BMI-1 and E2F3 expression and in a concomitant increase in E-cadherin levels.
  
  
  CONCLUSIONS
  miR-200c appears to control the EMT process through BMI-1 in bladder cancer cells, and it inhibits their proliferation through down-regulating E2F3. The targets of miR-200c include BMI-1 and E2F3, which are a novel regulator of EMT and a regulator of proliferation, respectively.
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. lychtm@gmail.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Liu
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. lcuzfy@163.com.
authors.forename: Mingning
authors.initials: M
authors.lastname: Qiu
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. ilppdameinv@163.com.
authors.forename: Guobin
authors.initials: G
authors.lastname: Tan
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. zijiliang@163.com.
authors.forename: Ziji
authors.initials: Z
authors.lastname: Liang
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. 465597410@qq.com.
authors.forename: Yue
authors.initials: Y
authors.lastname: Qin
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. chenlieqian@gmail.com.
authors.forename: Lieqian
authors.initials: L
authors.lastname: Chen
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. chenhege@hotmail.com.
authors.forename: Hege
authors.initials: H
authors.lastname: Chen
authors.affiliation: Laboratory of Urology, Guangdong Medical College, Zhanjiang, 524001, China. jianjunliulab@163.com.
authors.forename: Jianjun
authors.initials: J
authors.lastname: Liu
mesh_terms: D015703:Antigens, CD; D001483:Base Sequence; D015820:Cadherins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D049109:Cell Proliferation; D015536:Down-Regulation / Q000235:genetics*; D050699:E2F3 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D063150:Polycomb Repressive Complex 1 / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000473:pathology*
chemical_list: D015703:Antigens, CD; C069077:BMI1 protein, human; C513384:CDH1 protein, human; D015820:Cadherins; D050699:E2F3 Transcription Factor; C098319:E2F3 protein, human; C533896:MIRN200 microRNA, human; D035683:MicroRNAs; D063150:Polycomb Repressive Complex 1