Abstract
Recently, binding of specific protein 1 (Sp1) and cAMP response element binding protein (CREB) to a GC-rich element at -92/-62 has been identified as a critical step in gastrin-dependent regulation of the chromogranin A (CgA) gene in gastric epithelial cells. Here we demonstrate that binding of early growth response protein 1 (Egr-1) to the distal part of the -92/-62 site is also required for gastrin-dependent CgA transactivation. Gastrin elevated cellular and nuclear Egr-1 levels in a time-dependent manner and also increased Egr-1 binding to the CgA -92/-73 region. Disruption of this site reduced gastrin responsiveness without influencing basal promoter activity, while loss of Sp1 and/or CREB binding sites diminished basal and gastrin-stimulated CgA promoter activity. Ectopic Egr-1 overexpression potently stimulated the CgA promoter, whereas coexpression of Egr-1 with Sp1 and/or CREB resulted in additive effects. Functional analysis of Sp1-, Egr-1-, or CREB-specific promoter mutations in transfection studies confirmed the tripartite organization of the CgA -92/-62 element. Signaling studies revealed that MAPK kinase 1 (MEK1)/ERK1/2 cascades are critical for gastrin-dependent Egr-1 protein accumulation as well as Egr-1 binding to the CgA promoter. Our studies for the first time identify Egr-1 as a nuclear target of gastrin and show that functional interplay of Egr-1, Sp1, and CREB is indispensable for gastrin-dependent CgA transactivation in gastric epithelial cells.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Binding Sites
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Cell Nucleus / metabolism
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Chromogranin A
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Chromogranins / genetics*
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Cyclic AMP Response Element-Binding Protein / metabolism*
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DNA / chemistry
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DNA / metabolism
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DNA-Binding Proteins / metabolism*
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Early Growth Response Protein 1
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Enzyme Inhibitors / pharmacology
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Epithelial Cells / metabolism
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Flavonoids / pharmacology
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Gastrins / pharmacology*
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Humans
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Immediate-Early Proteins*
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Immunoblotting
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MAP Kinase Kinase 1
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Mitogen-Activated Protein Kinase 1 / metabolism
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinase Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinase Kinases / metabolism
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Mitogen-Activated Protein Kinases / metabolism
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Phosphorylation
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Promoter Regions, Genetic
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Protein Serine-Threonine Kinases / antagonists & inhibitors
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Protein Serine-Threonine Kinases / metabolism
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Response Elements
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Signal Transduction
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Stomach Neoplasms
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Tetradecanoylphorbol Acetate / pharmacology
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Transcription Factors / metabolism*
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Transcriptional Activation
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Transfection
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Tumor Cells, Cultured
Substances
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CHGA protein, human
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Chromogranin A
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Chromogranins
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Cyclic AMP Response Element-Binding Protein
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DNA-Binding Proteins
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EGR1 protein, human
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Early Growth Response Protein 1
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Enzyme Inhibitors
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Flavonoids
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Gastrins
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Immediate-Early Proteins
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Transcription Factors
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DNA
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Protein Serine-Threonine Kinases
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Mitogen-Activated Protein Kinase 1
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 1
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MAP2K1 protein, human
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Mitogen-Activated Protein Kinase Kinases
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Tetradecanoylphorbol Acetate
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2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one