Basal expression of I?B? is controlled by the mammalian transcriptional repressor RBP-J (CBF1) and its activator Notch1

Oakley, Fiona, Mann, Jelena, Ruddell, Richard G., Pickford, Jessica, Weinmaster, Gerry and Mann, Derek A. (2003) Basal expression of I?B? is controlled by the mammalian transcriptional repressor RBP-J (CBF1) and its activator Notch1. The Journal of Biological Chemistry, 278 (27), 24359-24370. (doi:10.1074/jbc.M211051200).

Abstract

By using the hepatic stellate cell (HSC) as a paradigm for cells that undergo long term re-programming of NF-?B-dependent transcription, we have determined a novel mechanism by which mammalian cells establish their basal NF-?B activity. Elevation of NF-?B activity during HSC activation is accompanied by induction of CBF1 expression and DNA binding activity. We show that the transcriptional repressor CBF1 interacts with a dual NF-?B/CBF1-binding site (?B2) in the I?B? promoter. Nucleotide substitutions that disrupt CBF1 binding to the ?B2 site result in an elevation of I?B? promoter activity and loss of responsiveness of the promoter to a transfected CBF1 reporter vector. Overexpression of CBF1 in COS1 cells was associated with markedly reduced I?B? protein expression and elevated NF-?B DNA binding activity. CBF1-induced repression of I?B? promoter activity was reversed in HSC transfected with the Notch1 intracellular domain (NICD). The ability of NICD to enhance I?B? gene transcription was confirmed in COS1 cells and was found to be dependent on an intact RAM domain of NICD that has been shown previously to help mediate the interaction of NICD with CBF1. One of the mechanisms by which NICD is thought to convert CBF1 into an activator of transcription is via the recruitment of transcriptional co-activators/histone acetylases to gene promoters. Co-transfection of HSC with NICD and p53 caused a diminution of I?B? promoter activity, by contrast overexpression of p300 enhanced IB? promoter function. Taken together, these data suggest that basal I?B? expression (and as a consequence NF-?B activity) is under the control of the various components of the CBF1/Notch signal transduction pathway.

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