title: MiR-291b-3p induces apoptosis in liver cell line NCTC1469 by reducing the level of RNA-binding protein HuR.
abstract: BACKGROUND
  There is increasing evidence that miRNAs are involved in cellular apoptosis. However, the specific role of miR-291b-3p in apoptosis has not been elucidated. In the present study, we investigated the effect of miR-291b-3p on NCTC1469 cell growth and apoptosis.
  
  
  METHODS
  Cell viability and apoptosis were examined in NCTC1469 cells transfected with miR-291b-3p mimics, inhibitor miRNA or negative control. Using computational miRNA target prediction databases, HuR was predicted as a target of miR-291b-3p. Luciferase assay, immunofluorescence and western blot were used to further explore the effects of miR-291b-3p on HuR expression. In addition, the effect of HuR on cell apoptosis was evaluated using a HuR-specific siRNA.
  
  
  RESULTS
  TNF-α-induced hepatocyte apoptosis was accompanied by enhanced expression of miR-291b-3p, suggesting that miR-291b-3p might contribute to the apoptotic process. Follow-up experiments showed that upregulation of miR-291b-3p decreased cell viability and induced NCTC1469 cell apoptosis. Additionally, similar to the activity of miR-519, which is another member of the same miRNA family, miR-291b-3p suppressed HuR translation through binding to the HuR coding region (CR). We further showed that the downregulation of HuR expression by miR-291b-3p was accompanied by reduced Bcl-2 expression. Moreover, knockdown of HuR also impaired Bcl-2 expression and increased the ratio of Bax/Bcl-2. More significantly, downregulation of miR-291b-3p failed to increase Bcl2 expression in NCTC1469 cells that were co-transfected with siRNA-HuR. Finally, inhibition of miR-291b-3p led to reduced apoptosis, while knockdown of HuR by siRNA promoted apoptosis, even in NCTC1469 cells that were co-transfected with the miR-291b-3p inhibitor.
  
  
  CONCLUSIONS
  The current data suggested that miR-291b-3p contributed to NCTC1469 cell apoptosis by regulating the expression of HuR, which in turn increased Bcl-2 stability.
authors.affiliation: Peking University Fifth School of Clinical Medicine, Beijing Hospital, Beijing, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Guo
authors.forename: Meng
authors.initials: M
authors.lastname: Li
authors.forename: Xiangyu
authors.initials: X
authors.lastname: Meng
authors.forename: JingZhe
authors.initials: J
authors.lastname: Sui
authors.forename: Lin
authors.initials: L
authors.lastname: Dou
authors.forename: Weiqing
authors.initials: W
authors.lastname: Tang
authors.forename: Xiuqing
authors.initials: X
authors.lastname: Huang
authors.forename: Yong
authors.initials: Y
authors.lastname: Man
authors.forename: Shu
authors.initials: S
authors.lastname: Wang
authors.forename: Jian
authors.initials: J
authors.lastname: Li
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002460:Cell Line; D000067780:ELAV-Like Protein 1 / Q000096:biosynthesis* / Q000235:genetics; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D008297:Male; D051379:Mice; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D014176:Protein Biosynthesis / Q000502:physiology*; D055550:Protein Stability; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000502:physiology*
chemical_list: D000067780:ELAV-Like Protein 1; C000593997:Elavl1 protein, mouse; D035683:MicroRNAs; D019253:Proto-Oncogene Proteins c-bcl-2; C052617:Bcl2 protein, mouse