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Molecular Pharmacology

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Research ArticleArticle

Both Inducible and Constitutive Activator Protein-1-Like Transcription Factors Are Used for Transcriptional Activation of the Galanin Gene by Different First and Second Messenger Pathways

Youssef Anouar, Hyeon-Woo Lee and Lee E. Eiden
Molecular Pharmacology July 1999, 56 (1) 162-169; DOI: https://doi.org/10.1124/mol.56.1.162
Youssef Anouar
Section on Molecular Neuroscience, Laboratory of Cellular and Molecular Regulation, National Institute of Mental Health, Bethesda, Maryland
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Hyeon-Woo Lee
Section on Molecular Neuroscience, Laboratory of Cellular and Molecular Regulation, National Institute of Mental Health, Bethesda, Maryland
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Lee E. Eiden
Section on Molecular Neuroscience, Laboratory of Cellular and Molecular Regulation, National Institute of Mental Health, Bethesda, Maryland
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Abstract

We investigated trans-acting factors mediating galanin (GAL) gene activation by protein kinase-dependent signal transduction pathways in chromaffin cells. GAL mRNA up-regulation via the protein kinase A (PKA) pathway (25 μM forskolin) required new protein synthesis. Stimulation via protein kinase C (0.1 μM phorbol myristate acetate) did not. The involvement of activator protein-1(AP-1) and cAMP response element-binding protein (CREB) in serine/threonine protein kinase activation of GAL gene transcription was assessed. Cotransfection of a GAL reporter gene along with expression plasmids encoding c-Jun plus c-Fos, or the catalytic subunit of PKA (PKAβ), resulted in a 4- to 8-fold enhancement of GAL reporter gene transcription. Transcriptional activation required the galanin 12-O-tetradecanoylphorbol-13-acetate (phorbol-12-myristate-13-acetate) response element (GTRE) octamer sequence (TGACGCGG) in the proximal enhancer of the GAL gene, previously shown to confer phorbol ester responsiveness in chromaffin cells. CREB coexpression did not stimulate GAL gene transcription or increase transcriptional activation by PKAβ. The GTRE preferentially bound in vitro synthesized Jun and Fos-Jun, compared with CREB, in electrophoretic mobility shift assays. The GTRE preference for binding AP-1-immunoreactive protein compared with CREB was even more pronounced in chromaffin cell nuclear extracts, in which the majority of GTRE-bound protein in electrophoretic mobility shift assays was supershifted with anti-Fos and anti-Jun antibodies. Thus, GAL gene regulation mediated by protein kinase activation appears to involve both constitutively expressed and inducible AP-1-related proteins. Elevated potassium stimulation of GAL mRNA was completely blocked, but pituitary adenylyl cyclase-activating polypeptide and histamine stimulations were only partially blocked, by cycloheximide. Both inducible and constitutive pathways are therefore used by physiologically relevant first messengers that stimulate GAL biosynthesis in vivo.

Footnotes

    • Received November 23, 1999.
    • Accepted March 15, 1999.
  • Send reprint requests to: Lee E. Eiden, Ph.D., Building 36, Room 2A-11, NIMH, NIH, 9000 Rockville Pike, Bethesda, MD 20892. E-mail:eiden{at}codon.nih.gov

  • ↵1 Present address: Unité Institut National de la Santé et de la Recherche Médicale U 413, Universitéde Rouen, Laboratoire de Neuroendocrinologie Cellulaire et Moléculaire, Institut Fédératif de Recherches Multidisciplinaires sur les Peptides, 76821 Mont-Saint-Aignan Cedex, France.

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Molecular Pharmacology: 56 (1)
Molecular Pharmacology
Vol. 56, Issue 1
1 Jul 1999
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Research ArticleArticle

Both Inducible and Constitutive Activator Protein-1-Like Transcription Factors Are Used for Transcriptional Activation of the Galanin Gene by Different First and Second Messenger Pathways

Youssef Anouar, Hyeon-Woo Lee and Lee E. Eiden
Molecular Pharmacology July 1, 1999, 56 (1) 162-169; DOI: https://doi.org/10.1124/mol.56.1.162

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Research ArticleArticle

Both Inducible and Constitutive Activator Protein-1-Like Transcription Factors Are Used for Transcriptional Activation of the Galanin Gene by Different First and Second Messenger Pathways

Youssef Anouar, Hyeon-Woo Lee and Lee E. Eiden
Molecular Pharmacology July 1, 1999, 56 (1) 162-169; DOI: https://doi.org/10.1124/mol.56.1.162
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