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Title: Cyclic adenosine 3',5'-monophosphate (cAMP) enhances cAMP-responsive element binding (CREB) protein phosphorylation and phospho-CREB interaction with the mouse steroidogenic acute regulatory protein gene promoter.

Authors: Clem, Brian F; Hudson, Elizabeth A; Clark, Barbara J

Published In Endocrinology, (2005 Mar)

Abstract: Steroidogenic acute regulatory protein (StAR) transcription is regulated through cAMP-protein kinase A-dependent mechanisms that involve multiple transcription factors including the cAMP-responsive element binding protein (CREB) family members. Classically, binding of phosphorylated CREB to cis-acting cAMP-responsive elements (5'-TGACGTCA-3') within target gene promoters leads to recruitment of the coactivator CREB binding protein (CBP). Herein we examined the extent of CREB family member phosphorylation on protein-DNA interactions and CBP recruitment with the StAR promoter. Immunoblot analysis revealed that CREB, cAMP-responsive element modulator (CREM), and activating transcription factor (ATF)-1 are expressed in MA-10 mouse Leydig tumor cells, yet only CREB and ATF-1 are phosphorylated. (Bu)2cAMP treatment of MA-10 cells increased CREB phosphorylation approximately 2.3-fold within 30 min but did not change total nuclear CREB expression levels. Using DNA-affinity chromatography, we now show that CREB and ATF-1, but not CREM, interact with the StAR promoter, and this interaction is dependent on the activator protein-1 (AP-1) cis-acting element within the cAMP-responsive region. In addition, (Bu)2cAMP-treatment increased phosphorylated CREB (P-CREB) association with the StAR promoter but did not influence total CREB interaction. In vivo chromatin immunoprecipitation assays demonstrated CREB binding to the StAR proximal promoter is independent of (Bu)2cAMP-treatment, confirming our in vitro analysis. However, (Bu)2cAMP-treatment increased P-CREB and CBP interaction with the StAR promoter, demonstrating for the first time the physical role of P-CREB:DNA interactions in CBP recruitment to the StAR proximal promoter.

PubMed ID: 15550512 Exiting the NIEHS site

MeSH Terms: Acetylation; Activating Transcription Factor 1; Animals; CREB-Binding Protein; Cell Nucleus/metabolism; Chromatin Immunoprecipitation; Chromatography; Cyclic AMP Response Element-Binding Protein/metabolism*; Cyclic AMP/metabolism*; DNA-Binding Proteins/metabolism; DNA/chemistry; Electrophoresis, Polyacrylamide Gel; Histones/metabolism; Immunoblotting; Leydig Cells; Male; Mice; Nuclear Proteins/metabolism*; Phosphoproteins/genetics*; Phosphorylation; Promoter Regions, Genetic; Signal Transduction; Time Factors; Trans-Activators/metabolism*; Transcription Factors/metabolism

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