Poster Presentation 30th Lorne Cancer Conference 2018

Investigating the in vivo role of the Ehf transcription factor in intestinal epithelial differentiation and tumorigenesis (#248)

Camilla Reehorst 1 2 , Rebecca Nightingale 1 , Ian Luk 1 3 , Amardeep Dhillon 1 2 , John Mariadason 1 2
  1. Olivia Newton-John Cancer Research Institute, Heidelberg, VICTORIA, Australia
  2. School of Cancer Medicine, La Trobe University, Bundoora, VIC, Australia
  3. Department of Medicine, The University of Melbourne , Parkville, VIC, Australia

Objective
Poor tumour differentiation status is associated with worse patient outcome in colorectal cancer. However, the molecular basis for this loss of differentiation is not well understood.  We have found that the transcription factor Ets homologous factor (EHF) is highly expressed in the normal human and mouse colonic epithelium and is down-regulated in poorly-differentiated human colorectal cancer cell lines. The aim of this study was to investigate the role of murine Ehf in regulating normal colonic epithelial cell differentiation in vivo and how loss of Ehf impacts Apc-initiated tumourigenesis. 

Methods
A novel mouse model was generated in which the Ets DNA binding domain (exon 8) of Ehf was flanked by loxP sites (Ehflox/lox).  Ehflox/lox mice were crossed with VillinCreERT2 mice and treated with tamoxifen to induce intestinal-specific inactivation of Ehf. The effect of Ehf deletion on intestinal cell proliferation and differentiation was assessed by qRT-PCR and immunohistochemistry. The impact of Ehf deletion on colonic adenoma formation and survival was determined by crossing Ehflox/lox mice with Apclox/lox; Cdx2CreERT2 mice and treat with tamoxifen to induce colon-specific deletion of Ehf and Apc.

Results
Targeted inactivation of Ehf in the intestinal epithelium following tamoxifen administration was confirmed by qRT-PCR and genotyping.  Ehf inactivation in the intestinal epithelium had minimal effect on survival and overall health of the animals. Furthermore, loss of Ehf did not markedly affect intestinal cell proliferation and expression of markers of absorptive, enteroendocrine or Paneth cell differentiation. Comparatively, inactivation of Ehf increased the number of goblet cells in the colonic epithelium, and caused marked transcriptional changes including reduced expression of cholesterol transport genes. Finally, Ehf inactivation in the colonic epithelium had minimal effect on Apc-initiated colonic adenoma formation, histology and survival.

Conclusion
Inactivation of Ehf alone in the intestinal epithelium has minimal impact on intestinal cell homeostasis and adenoma formation, but does result in significant gene expression changes. Recent in vitro studies in our laboratory demonstrate that Ehf cooperates with Cdx1 to regulate colonic epithelial cell differentiation. Work is underway to investigate the impact of combined Cdx1 and Ehf deletion on homeostasis of the colonic epithelium and Apc-initiated tumourigenesis.