Suppression of beta-casein gene expression by inhibition of protein synthesis in mouse mammary epithelial cells is associated with stimulation of NF-kappaB activity and blockage of prolactin-Stat5 signaling.
The protein synthesis inhibitor cycloheximide (Chx) suppresses prolactin-induced beta-casein gene expression in the mammary epithelial cell line COMMA-D. As the mechanism underlying this effect is unclear, the effects of protein synthesis inhibitors on interactions of transcription factors with the beta-casein promoter were examined. Suppression of prolactin-induced beta-casein gene expression occurred in both COMMA-D cells and primary mammary cell cultures with as little as 2 h protein synthesis inhibition. This was associated with changes in transcription factors interacting at a response element in the proximal region of the rat beta-casein promoter. Inhibition of protein synthesis was associated with NF-kappaB binding at a site immediately 3' to the Stat5-binding site at position 97-89 of the beta-casein promoter, suppression of Stat5 DNA-binding activity, and inhibition of Stat5 tyrosine phosphorylation. Treatment with the NF-kappaB inhibitor parthenolide failed to restore prolactin responsiveness. These results show that protein synthesis inhibition is associated with both blockage of prolactin-Stat5 signaling and NF-kappaB binding to the beta-casein promoter, but that the latter is not necessary for the suppression of beta-casein expression.
Beaton
Angela
A
AgResearch, Ruakura Research Centre, Private Bag 3123, Hamilton, New Zealand.
Broadhurst
Marita K
MK
Wilkins
Richard J
RJ
Wheeler
Thomas T
TT
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002364:Caseins / Q000235:genetics*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000502:physiology*; D004847:Epithelial Cells / Q000166:cytology / Q000201:enzymology / Q000502:physiology*; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D008321:Mammary Glands, Animal / Q000166:cytology / Q000201:enzymology / Q000502:physiology*; D051379:Mice; D008894:Milk Proteins; D016328:NF-kappa B / Q000378:metabolism*; D011388:Prolactin / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D050799:STAT5 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D015534:Trans-Activators / Q000502:physiology*
D002364:Caseins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D016328:NF-kappa B; D011500:Protein Synthesis Inhibitors; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D011388:Prolactin; D003513:Cycloheximide