Hypoxia-inducible factors modulate the stemness and malignancy of colon cancer cells by playing opposite roles in canonical Wnt signaling

PLoS One. 2014 Nov 14;9(11):e112580. doi: 10.1371/journal.pone.0112580. eCollection 2014.

Abstract

This study examined the role played by hypoxia-inducible factors (HIFs) in malignant phenotype maintenance and canonical Wnt signaling. Under normoxia, we determined that both HIF-1α and HIF-2α are expressed in human colon cancer cells but not in their non-malignant counterparts. The stable knockdown of HIF-1α or HIF-2α expression induced negative effects on the malignant phenotype of colon cancer cells, with lactate production, the rate of apoptosis, migration, CXCR4-mediated chemotaxis, and tumorigenic activity all being significantly affected by HIF knockdown and with HIF-1α depletion exerting greater effects. Knockdown of these two HIF transcripts induced different and even opposite effects on β-catenin transcriptional activity in colon cancer cells with different genetic Wnt signaling pathways. In SW480 cells, HIF-2α knockdown did not affect β-catenin levels, increasing the transcriptional activity of β-catenin by inducing its nuclear accumulation, whereas HIF-1α silencing negatively affected the stability and transcriptional activity of β-catenin, inducing its exit from the nuclei and its recruitment to the cell membrane by E-cadherin. In addition, although HIF-1α depletion induced a reversal of the epithelial-to-mesenchymal transition (EMT), HIF-2α silencing altered the expression of the stem cell markers CD44, Oct4, and CD24 and of the differentiation marker CK20 in the opposite direction as HIF-1α silencing. Remarkably, HIF-2α knockdown also enhanced β-catenin transcriptional activity under hypoxia in cells that displayed normal Wnt signaling, suggesting that the gene negatively modulates canonical Wnt signaling in colon cancer cells. Taken together, our results indicate that HIFs play opposing roles in canonical Wnt signaling and are essential for the stemness and malignancy maintenance of colon cancer cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Basic Helix-Loop-Helix Transcription Factors / genetics
  • Basic Helix-Loop-Helix Transcription Factors / metabolism*
  • Blotting, Western
  • Cell Line, Tumor
  • Colonic Neoplasms / metabolism*
  • Colonic Neoplasms / pathology*
  • Flow Cytometry
  • Gene Expression Regulation, Neoplastic / physiology*
  • Gene Knockdown Techniques
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit / genetics
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism*
  • Immunoprecipitation
  • Wnt Signaling Pathway / physiology*
  • beta Catenin / metabolism

Substances

  • Basic Helix-Loop-Helix Transcription Factors
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • beta Catenin
  • endothelial PAS domain-containing protein 1

Grant support

This research was supported by grants from Universidad Nacional Autónoma de México (DGAPA-UNAM IN226111 and IN215514) and Consejo Nacional de Ciencia y Tecnología (CONACYT CB2011-151731). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.