Down-regulation of microRNA-9 leads to activation of IL-6/Jak/STAT3 pathway through directly targeting IL-6 in HeLa cell

Mol Carcinog. 2016 May;55(5):732-42. doi: 10.1002/mc.22317. Epub 2015 Mar 25.


MicroRNA-9 (miR-9) presents to exert distinct and even opposite functions in different kinds of tumors through targeting different cellular genes. However, its role in cervical adenocarcinoma remains uncertain. Here, we report that miR-9 is down-regulated in cervical adenocarcinoma due to its frequent promoter-hypermethylation and exerts its tumor suppressor role through inhibiting several novel target genes, including interleukin-6 (IL-6). The promoters of miR-9 precursors (mir-9-1, -2, and -3) were hypermethylated in cervical adenocarcinoma tissues. Demethylation treatment of HeLa dramatically increased the expression of mature miR-9. Both in vitro and in vivo functional experiments confirmed that miR-9 can inhibit the proliferation, migration, and malignant transformation abilities of HeLa cells. Bioinformatics methods and array-based RNA expression profiles were used to screen the downstream target genes of miR-9. Dual-luciferase reporting assay, real-time qPCR, and ELISA or Western blot confirmed four genes (CKAP2, HSPC159, IL-6, and TC10) to be novel direct target genes of miR-9. Pathway annotation analysis of the differently expressed genes (DEGs) induced by ectopic miR-9 expression revealed the enrichment in Jak/STAT3 pathway, which is one of the downstream pathways of IL-6. Ectopic expression of miR-9 in HeLa inhibited Jak/STAT3 signaling activity. Moreover, such effect could be partially reversed by the addition of exogenous IL-6. In conclusion, our results here present a tumor suppressor potential of miR-9 in cervical adenocarcinoma for the first time and suggest that miR-9 could repress tumorigenesis through inhibiting the activity of IL-6/Jak/STAT3 pathway.

Keywords: HeLa; Jak/STAT3 signaling pathway; cervical adenocarcinoma; hypermethylation; interleukin-6; microRNA-9-1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions
  • Animals
  • Cell Proliferation
  • DNA Methylation
  • Down-Regulation*
  • Female
  • Gene Expression Regulation, Neoplastic
  • HeLa Cells
  • Humans
  • Interleukin-6 / genetics*
  • Interleukin-6 / metabolism
  • Janus Kinases / genetics
  • Janus Kinases / metabolism
  • Mice
  • MicroRNAs / genetics*
  • MicroRNAs / metabolism*
  • Neoplasm Transplantation
  • Promoter Regions, Genetic
  • STAT3 Transcription Factor / genetics
  • STAT3 Transcription Factor / metabolism
  • Signal Transduction
  • Uterine Cervical Neoplasms / genetics*
  • Uterine Cervical Neoplasms / metabolism
  • Uterine Cervical Neoplasms / pathology


  • 3' Untranslated Regions
  • IL6 protein, human
  • Interleukin-6
  • MIRN92 microRNA, human
  • MicroRNAs
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • Janus Kinases