Activation of protein kinase Calpha by EPAC1 is required for the ERK- and CCAAT/enhancer-binding protein beta-dependent induction of the SOCS-3 gene by cyclic AMP in COS1 cells

Gillian Borland, Rebecca J. Bird, Timothy M. Palmer, Stephen J Yarwood

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    We recently found that induction of the anti-inflammatory SOCS-3 gene by cyclic AMP occurs through novel cyclic AMP-dependent protein kinase-independent mechanisms involving activation of CCAAT/enhancer-binding protein (C/EBP) transcription factors, notably C/EBPbeta, by the cyclic AMP GEF EPAC1 and the Rap1 GTPase. In this study we show that down-regulation of phospholipase (PL) Cepsilon with small interfering RNA or blockade of PLC activity with chemical inhibitors ablates exchange protein directly activated by cyclic AMP (EPAC)-dependent induction of SOCS-3 in COS1 cells. Consistent with this, stimulation of cells with 1-oleoyl-2-acetyl-sn-glycerol and phorbol 12-myristate 13-acetate, both cell-permeable analogues of the PLC product diacylglycerol, are sufficient to induce SOCS-3 expression in a Ca2+-dependent manner. Moreover, the diacylglycerol- and Ca2+-dependent protein kinase C (PKC) isoform PKCalpha becomes activated following cyclic AMP elevation or EPAC stimulation. Conversely, down-regulation of PKC activity with chemical inhibitors or small interfering RNA-mediated depletion of PKCalpha or -delta blocks EPAC-dependent SOCS-3 induction. Using the MEK inhibitor U0126, we found that activation of ERK MAPKs is essential for SOCS-3 induction by either cyclic AMP or PKC. C/EBPbeta is known to be phosphorylated and activated by ERK. Accordingly, we found ERK activation to be essential for cyclic AMP-dependent C/EBP activation and C/EBPbeta-dependent SOCS-3 induction by cyclic AMP and PKC. Moreover, overexpression of a mutant form of C/EBPbeta (T235A), which lacks the ERK phosphorylation site, blocks SOCS-3 induction by cyclic AMP and PKC in a dominant-negative manner. Together, these results indicate that EPAC mediates novel regulatory cross-talk between the cyclic AMP and PKC signaling pathways leading to ERK- and C/EBPbeta-dependent induction of the SOCS-3 gene.

    Original languageEnglish
    Pages (from-to)17391-17403
    Number of pages13
    JournalJournal of Biological Chemistry
    Issue number26
    Publication statusPublished - 26 Jun 2009


    • Animals
    • Blotting, Western
    • CCAAT-Enhancer-Binding Protein-beta
    • COS Cells
    • Cercopithecus aethiops
    • Cyclic AMP
    • Extracellular Signal-Regulated MAP Kinases
    • Gene Expression
    • Guanine Nucleotide Exchange Factors
    • Humans
    • Luciferases
    • Phosphoinositide Phospholipase C
    • Phosphorylation
    • Protein Kinase C-alpha
    • RNA, Small Interfering
    • Signal Transduction
    • Suppressor of Cytokine Signaling Proteins
    • Telomere-Binding Proteins
    • Tetradecanoylphorbol Acetate


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