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  • Binding of C/EBPbeta to the C-reactive protein (CRP) promoter in Hep3B cells is associated with transcription of CRP mRNA.

Binding of C/EBPbeta to the C-reactive protein (CRP) promoter in Hep3B cells is associated with transcription of CRP mRNA.

Journal of immunology (Baltimore, Md. : 1950) (2008-08-08)
Duprane Pedaci Young, Irving Kushner, David Samols

Expression of the acute phase protein C-reactive protein (CRP) is tightly regulated in hepatocytes. Although very little CRP mRNA is transcribed normally, inflammatory stimuli are followed by a dramatic increase in mRNA synthesis and accumulation. IL-6 and IL-1beta are believed to be the major cytokines responsible for induction of CRP and other acute phase proteins. Our previous studies, using transient transfection and EMSA experiments, implicated involvement of the transcription factors C/EBPbeta, STAT3, Rel p50, and c-Rel in CRP induction. In the current study we used chromatin immunoprecipitation assays to determine the kinetics of transcription factor occupancy of these transcription factors on the endogenous CRP promoter. All of these transcription factors were found bound to the endogenous CRP promoter in the absence of cytokines, but cytokine treatment markedly increased binding of only C/EBPbeta. In addition, c-Rel and TATA box-binding protein (TBP) appeared to occupy the promoter in parallel in the presence of cytokines. In the absence of cytokines, CRP mRNA accumulation was not measurable but began to increase by 3 h after exposure of cells to IL-1beta plus IL-6, peaking at 12 h with secondary peaks at 18 and 24 h. The secondary peaks in mRNA expression paralleled the pattern of binding of c-Rel and TBP to the CRP promoter. We conclude that the CRP promoter has a low level of transcription factor occupancy in the absence of cytokines and induction occurs with binding of C/EBP, and that c-Rel and TBP are important for modulating CRP expression.

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Deoxyribonucleic acid, single stranded from salmon testes, For hybridization
Goat Anti-Rabbit IgG Antibody, HRP-conjugate, 1 mg/mL, Upstate®
Protein A Agarose/Salmon Sperm DNA, 2.5 mL, Recombinant protein A covalently bound to agarose beads for use in chromatin immunoprecipitations (ChIP assays). Sonicated salmon sperm DNA is included to block non-specific DNA binding sites on agarose beads. Every lot qualified using an Acetyl Histone H4 and H3 ChIP assay.