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Increased GADD gene expression in human colon epithelial cells exposed to deoxycholate
David W Scott1, Sophia Mutamba, Robin G Hopkins
1Cellular and Molecular Nutrition Research Laboratory, Graduate Program in Nutrition, University of North Carolina at Greensboro, Greensboro, North Carolina 27403-6170, USA.
Journal of Cellular Physiology
|August 19, 2004
Summary
Secondary bile acids like deoxycholate (DOC) can stress colon cells. This study found DOC upregulates growth arrest and DNA damage-inducible (GADD) genes, particularly GADD153, in human colon cells, independent of p53 status.
Area of Science:
- Molecular biology
- Cellular stress response
- Gastrointestinal physiology
Background:
- Colonic epithelial cells face high secondary bile acid concentrations.
- Secondary bile acids can induce cellular stress and activate stress-response genes.
- Growth arrest and DNA damage-inducible (GADD) genes are key players in cellular stress pathways.
Purpose of the Study:
- To investigate the in vitro effect of deoxycholate (DOC) on GADD gene expression in human colonic epithelial cells.
- To determine if p53 status influences DOC-induced GADD gene upregulation.
- To elucidate the molecular mechanisms underlying DOC-induced GADD153 expression.
Main Methods:
- DNA macroarray screening of stress/apoptosis-related genes in DOC-treated HCT-116 colonocytes.
- Gene-specific relative RT-PCR to confirm GADD gene expression changes.
- Experiments using actinomycin-D, antioxidants, MAPK inhibitors, anisomycin, and cycloheximide to study GADD153 regulation.
Main Results:
- Deoxycholate (DOC) significantly upregulated GADD45, GADD34, and most notably GADD153 mRNA expression in HCT-116 colonocytes.
- DOC-induced upregulation of GADD34, GADD45, and GADD153 mRNA occurred independently of p53 status, as shown in HCT-15 cells.
- DOC-induced GADD153 mRNA upregulation was transcriptional and not mediated by reactive oxygen species or MAPK signaling, while protein expression involved translation of pre-existing mRNA.
Conclusions:
- Deoxycholate (DOC) is a potent inducer of GADD gene expression in human colonic epithelial cells.
- The upregulation of GADD genes, particularly GADD153, by DOC is a p53-independent stress response.
- DOC-induced GADD153 expression involves transcriptional activation and post-transcriptional regulation of protein synthesis.