A novel RBP-J kappa-dependent switch from C/EBP beta to C/EBP zeta at the C/EBP binding site on the C-reactive protein promoter.
Regulation of basal and cytokine (IL-6 and IL-1beta)-induced expression of C-reactive protein (CRP) in human hepatoma Hep3B cells occurs during transcription. A critical transcriptional regulatory element on the CRP promoter is a C/EBP binding site overlapping a NF-kappaB p50 binding site. In response to IL-6, C/EBPbeta and p50 occupy the C/EBP-p50 site on the CRP promoter. The aim of this study was to identify the transcription factors occupying the C/EBP-p50 site in the absence of C/EBPbeta. Accordingly, we treated Hep3B nuclear extract with a C/EBP-binding consensus oligonucleotide to generate an extract lacking active C/EBPbeta. Such treated nuclei contain only C/EBPzeta (also known as CHOP10 and GADD153) because the C/EBP-binding consensus oligonucleotide binds to all C/EBP family proteins except C/EBPzeta. EMSA using this extract revealed formation of a C/EBPzeta-containing complex at the C/EBP-p50 site on the CRP promoter. This complex also contained RBP-Jkappa, a transcription factor known to interact with kappaB sites. RBP-Jkappa was required for the formation of C/EBPzeta-containing complex. The RBP-Jkappa-dependent C/EBPzeta-containing complexes were formed at the C/EBP-p50 site on the CRP promoter in the nuclei of primary human hepatocytes also. In luciferase transactivation assays, overexpressed C/EBPzeta abolished both C/EBPbeta-induced and (IL-6 + IL-1beta)-induced CRP promoter-driven luciferase expression. These results indicate that under basal conditions, C/EBPzeta occupies the C/EBP site, an action that requires RBP-Jkappa. Under induced conditions, C/EBPzeta is replaced by C/EBPbeta and p50. We conclude that the switch between C/EBPbeta and C/EBPzeta participates in regulating CRP transcription. This process uses a novel phenomenon, that is, the incorporation of RBP-Jkappa into C/EBPzeta complexes solely to support the binding of C/EBPzeta to the C/EBP site.
Singh
Prem Prakash
PP
Department of Pharmacology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, USA.
Voleti
Bhavya
B
Agrawal
Alok
A
D001665:Binding Sites / Q000235:genetics / Q000276:immunology; D002097:C-Reactive Protein / Q000235:genetics* / Q000378:metabolism*; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000172:deficiency / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D022762:CCAAT-Enhancer-Binding Proteins / Q000096:biosynthesis / Q000378:metabolism* / Q000502:physiology; D045744:Cell Line, Tumor; D051858:Forkhead Transcription Factors / Q000378:metabolism; D051537:Hepatocyte Nuclear Factor 1 / Q000378:metabolism; D006801:Humans; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein / Q000378:metabolism / Q000502:physiology*; D007134:Immunoglobulin Switch Region / Q000235:genetics / Q000276:immunology*; D052002:NF-kappa B p50 Subunit / Q000378:metabolism; D050811:Octamer Transcription Factor-1 / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000276:immunology*; D051742:Transcription Factor CHOP / Q000096:biosynthesis / Q000378:metabolism* / Q000502:physiology; D014158:Transcription, Genetic
D022782:CCAAT-Enhancer-Binding Protein-beta; D022762:CCAAT-Enhancer-Binding Proteins; C497249:DDIT3 protein, human; C477894:FOXI1 protein, human; D051858:Forkhead Transcription Factors; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D052002:NF-kappa B p50 Subunit; D050811:Octamer Transcription Factor-1; C497570:RBPJ protein, human; D051537:Hepatocyte Nuclear Factor 1; D051742:Transcription Factor CHOP; D002097:C-Reactive Protein