Proteasome inhibitors activate the transcription factors C/EBP-beta and delta in human intestinal epithelial cells.
In recent studies, induction of the heat shock response by hyperthermia upregulated the expression and DNA binding activity of the transcription factor C/EBP. This is an important observation because it may at least in part explain why the heat shock response upregulates IL-6 production in the intestinal mucosa and in the enterocyte. A novel method to induce the heat shock response is proteasome inhibition. The influence of this treatment on the expression and DNA binding activity of C/EBP is not known. We treated cultured Caco-2 cells, a human intestinal epithelial cell line, with one of the proteasome inhibitors, MG-132 or lactacystin, and measured C/EBP-beta and delta DNA binding activity by electrophoretic mobility shift assay and supershift analysis. In addition, nuclear levels of C/EBP-beta and delta protein were determined by Western blot analysis. Treatment of the cells with the proteasome inhibitors resulted in increased cellular levels of heat shock protein 72, consistent with induction of the heat shock response. Treatment also resulted in increased DNA binding activity and nuclear protein levels of C/EBP-beta and delta. The effects of the proteasome inhibitors on C/EBP were inhibited by treating the cells with quercetin, a substance known to block the heat shock response. The results suggest that proteasome inhibition activates the transcription factors C/EBP-beta and delta in human intestinal epithelial cells and that this response, at least in part, is caused by induction of the heat shock response. The observations are important because they provide support for a novel method to influence gene activation in the enterocyte.
Hungness
Eric S
ES
Department of Surgery, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267, USA.
Robb
Bruce W
BW
Luo
Guang-ju
GJ
Pritts
Timothy A
TA
Hershko
Dan D
DD
Hasselgren
Per-Olof
PO
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D015153:Blotting, Western; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism*; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D018938:Caco-2 Cells; D002467:Cell Nucleus / Q000378:metabolism; D003546:Cysteine Endopeptidases; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D020895:Enterocytes / Q000378:metabolism; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000378:metabolism*; D050884:HSP72 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007976:Leupeptins / Q000494:pharmacology; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors*; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D020033:Protein Isoforms; D011794:Quercetin / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors; D014157:Transcription Factors
D022782:CCAAT-Enhancer-Binding Protein-beta; D022762:CCAAT-Enhancer-Binding Proteins; C497252:CEBPD protein, human; D015853:Cysteine Proteinase Inhibitors; D050884:HSP72 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D007976:Leupeptins; D009097:Multienzyme Complexes; D020033:Protein Isoforms; D014157:Transcription Factors; C067713:lactacystin; D051743:CCAAT-Enhancer-Binding Protein-delta; D004247:DNA; D011794:Quercetin; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D000111:Acetylcysteine