Knockdown of β-Catenin through shRNA cause a reversal of EMT and metastatic phenotypes induced by HIF-1α

Cancer Invest. 2011 Jul;29(6):377-82. doi: 10.3109/07357907.2010.512595.


Objective: Wnt/β-catenin signaling pathway regulates pattern formation during embryogenesis as well as tumor progression. Numbers of studies suggest that this signaling pathway may play an important role in Epithelial-Mesenchymal transition (EMT), however, there was no evidence that Wnt/β-catenin signaling pathway directly controlled the EMT occurrence. Our previous research has successfully proved that overexpression of hypoxia inducible factor-1α (HIF-1α) could induce EMT in LNCaP cells, but not in PC-3. Consistently, the expression of β-catenin protein increased in LNCaP/HIF-1α cells, but not in PC-3/HIF-1α. This study mainly aimed at exploring the essentiality and importance of Wnt/β-catenin signaling pathway in HIF-1α-induced EMT.

Methods: Human prostate cancer cells (LNCaP) were stably transfected by recombinant plasmid pcDNA3.1(-)/HIF-1α. The positive clones were selected by G418 and confirmed through western blot analysis, reverse transcription-polymerase chain reaction (RT-PCR), and indirect immunofluoesence. Then LNCaP/HIF-1α was transiently transfected with β-catenin shRNA (shRNA1 and shRNA2) and negative shRNA (shRNA-scr). The epithelial markers, mesenchymal markers, and critical proteins in Wnt/β-catenin signaling pathway were separately detected by western blot analysis. Finally, the invasive potency of cells in different transfection group was examined by Matrgel transwell assay.

Result: We successfully established prostate cancer cell line LNCaP/HIF-1α and LNCaP/HIF-1α/β-catenin(-). LNCaP/HIF-1α displayed high expression of mesenchymal markers and low expression of epithelial markers. However, compared with LNCaP/HIF-1α, the epithelial marker E-cadherin was increased in LNCaP/HIF-1α/β-catenin(-), whereas the expression of mesenchymal marker N-cadherin, vimentin, MMP-2 were significantly decreased. Inhibition of Wnt signal activity through β-catenin shRNA cause a reversal of EMT induced by HIF-1α in human prostate cancer.

Conclusion: Overexpression of HIF-1α stimulates the invasion potency of human prostate carcinoma cells through EMT pathway and Wnt/β-catenin signaling pathway played a vital role in this process. Wnt/β-catenin signaling pathway might be a necessary endogenous signal that directly controlled the EMT occurrence induced by HIF-1α.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line, Tumor
  • Epithelial-Mesenchymal Transition*
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit / physiology*
  • Male
  • Neoplasm Invasiveness
  • Neoplasm Metastasis
  • Phenotype
  • Prostatic Neoplasms / pathology*
  • RNA, Small Interfering / genetics*
  • Signal Transduction / physiology
  • Wnt Proteins / physiology
  • beta Catenin / antagonists & inhibitors
  • beta Catenin / physiology*


  • CTNNB1 protein, human
  • HIF1A protein, human
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • RNA, Small Interfering
  • Wnt Proteins
  • beta Catenin