Characterization of Ca2+/Calmodulin-dependent protein kinase IV: Role in transcriptional regulation

Hervé Enslen, Peiqing Sun, Debra Brickey, Scott H. Soderling, Elizabeth Klamo, Thomas Soderling

    Research output: Contribution to journalArticle

    328 Citations (Scopus)

    Abstract

    We have characterized Ca2+/calmodulin-dependent protein kinase IV (CaM kinase IV), expressed using the baculovirus/Sf9 cell system, to assess its potential role in Ca2+-dependent transcriptional regulation. CaM kinase TV was strongly inhibited in vitro by KN-62, a specific CaM kinase inhibitor which suppresses Ca2+-dependent transcription of several genes, so we tested whether CaM kinase IV could stimulate transcription. Co-transfection of COS-1 cells by cDNA for CaM kinase IV gave 3-fold stimulation of a reporter gene expression, whereas co-transfection with CaM kinase II gave no transcriptional stimulation. Since this transcriptional response was mediated by phosphorylation of cAMP responsive element-binding protein (CREB), we determined the kinetics and site specificities of CaM kinases IV and II for phosphorylating CREB in vitro. CaM kinases IV and II and cAMP kinase (protein kinase A) all had similar Km values for CREB (1-5 μM), but the Vmax of CaM kinase IV was 40-fold lower than those of CaM kinase II and protein kinase A. Although all three kinases phosphorylated Ser133 in CREB, CaM kinase II also gave equal phosphorylation of a second site which was not Ser98. The two CREB phosphorylation sites were separately 32P-labeled, and the abilities of protein phosphatases 1, 2A, and 2B (calcineurin) to dephosphorylate them were tested. Our results show that all three phosphatases could dephosphorylate both sites, and calcineurin was a stronger catalyst for dephosphorylating site 1 (Ser133) than for site 2. These results indicate that CaM kinase TV may be important in Ca2+-dependent transcriptional regulation through phosphorylation of Ser133 in CREB. The fact that CaM kinase II phosphorylates another site in addition to Ser133 in CREB raises the possibility that this second phosphorylation site may account for the suppressed ability of CaM kinase II to enhance transcription through the CRE/CREB system. In addition multiple protein phosphatases, including calcineurin, may exert a modulatory effect on transcription depending on which site they dephosphorylate.

    Original languageEnglish (US)
    Pages (from-to)15520-15527
    Number of pages8
    JournalJournal of Biological Chemistry
    Volume269
    Issue number22
    StatePublished - Jun 3 1994

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    Calcium-Calmodulin-Dependent Protein Kinase Type 4
    Calcium-Calmodulin-Dependent Protein Kinase Type 2
    Carrier Proteins
    Phosphorylation
    Calcineurin
    Transcription
    Phosphotransferases
    KN 62
    Cyclic AMP-Dependent Protein Kinases
    Transfection
    Protein Phosphatase 1
    Sf9 Cells
    Protein Phosphatase 2
    Baculoviridae
    Phosphoprotein Phosphatases
    COS Cells
    Reporter Genes
    Phosphoric Monoester Hydrolases
    Gene expression
    Protein Kinases

    ASJC Scopus subject areas

    • Biochemistry

    Cite this

    Enslen, H., Sun, P., Brickey, D., Soderling, S. H., Klamo, E., & Soderling, T. (1994). Characterization of Ca2+/Calmodulin-dependent protein kinase IV: Role in transcriptional regulation. Journal of Biological Chemistry, 269(22), 15520-15527.

    Characterization of Ca2+/Calmodulin-dependent protein kinase IV : Role in transcriptional regulation. / Enslen, Hervé; Sun, Peiqing; Brickey, Debra; Soderling, Scott H.; Klamo, Elizabeth; Soderling, Thomas.

    In: Journal of Biological Chemistry, Vol. 269, No. 22, 03.06.1994, p. 15520-15527.

    Research output: Contribution to journalArticle

    Enslen, H, Sun, P, Brickey, D, Soderling, SH, Klamo, E & Soderling, T 1994, 'Characterization of Ca2+/Calmodulin-dependent protein kinase IV: Role in transcriptional regulation', Journal of Biological Chemistry, vol. 269, no. 22, pp. 15520-15527.
    Enslen H, Sun P, Brickey D, Soderling SH, Klamo E, Soderling T. Characterization of Ca2+/Calmodulin-dependent protein kinase IV: Role in transcriptional regulation. Journal of Biological Chemistry. 1994 Jun 3;269(22):15520-15527.
    Enslen, Hervé ; Sun, Peiqing ; Brickey, Debra ; Soderling, Scott H. ; Klamo, Elizabeth ; Soderling, Thomas. / Characterization of Ca2+/Calmodulin-dependent protein kinase IV : Role in transcriptional regulation. In: Journal of Biological Chemistry. 1994 ; Vol. 269, No. 22. pp. 15520-15527.
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