Regulation of hypoxia-inducible factor 1 in enterocytic cells.
BACKGROUND
Mucosal hypoxia due to intestinal hypoperfusion is characteristic of a number of clinical disorders. An early event in the adaptive response to cellular hypoxia is the binding of hypoxia-inducible factor 1 (HIF-1) to cis-acting regulatory sites in target genes.


METHODS
We characterized the expression of HIF-1 in transformed (Caco-2(BBe) and T84) and nontransformed human (FHs 74 Int) and rat (IEC-6) intestinal epithelial cell lines.


RESULTS
The electrophoretic mobility shift assay detected increased HIF-1 DNA-binding activity in each cell line within 2 h of hypoxia (1% O2). HIF-1 binding was maximal within 4 h and remained stable for 24 h. HIF-1 DNA-binding activity was maximal in the established IEC-6 cell line below 2% oxygen, but HIF-1 DNA-binding activity was not detectable above 0.5% O2 in the primary human FHs 74 Int cell line. The nonspecific protein kinase inhibitor genistein (200 microM) inhibited HIF-1 binding at 4 h. Transfection of Caco-2 cells with a wild-type, but not a mutant, HIF-1-dependent luciferase expression vector resulted in a fourfold induction of reporter gene expression during hypoxia.


CONCLUSIONS
In conclusion, HIF-1 regulates gene expression in enterocytes and an undefined phosphorylation event is important for O2 sensing.
Department of Surgery, Beth Israel Deaconess Medical Center and Harverd Medical School, Boston, MA 02215, USA.
Daniel J
DJ
Bertges
Sören
S
Berg
Mitchell P
MP
Fink
Russell L
RL
Delude
D000818:Animals; D018938:Caco-2 Cells; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D020895:Enterocytes / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007413:Intestinal Mucosa / Q000166:cytology; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D047428:Protein Kinase Inhibitors; D051381:Rats; D015398:Signal Transduction / Q000502:physiology; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; C497442:HIF1A protein, human; C497444:Hif1a protein, rat; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D014157:Transcription Factors; D019833:Genistein; D010100:Oxygen