Abstract
Notch-1 belongs to a family of transmembrane receptor proteins that direct the decisions as to various cell fates. After ligand binding, a proteolytic cleavage step occurs and the intracellular part of Notch-1, Notch-1-IC, translocates into the nucleus, where it targets the DNA binding protein RBP-J kappa/CBF1. RBP-J kappa mediates repression through recruitment of a histone deacetylase-containing complex. The Notch-1-IC/RBP-J kappa complex overcomes repression and activates the transcription of Notch target genes. We have identified a novel domain in Notch-1-IC, the EP domain, which is indispensable for full transcriptional activation. This transactivation domain is localized adjacent to the ankyrin repeats of Notch-1-IC. In cotransfection experiments, Notch-1-IC-mediated transcriptional activation was inhibited by E1A12S and p53, two proteins, which interfere with the function of the common coactivator p300. Protein-protein interaction assays demonstrated the association of Notch-1-IC and the CH3 region of p300. In addition, the interaction of mammalian Notch-1-IC with p300 was destabilized after deletion of the EP domain of Notch-1-IC. Based on physical interaction with Notch-1-IC and coactivator functions of p300, we propose a model for Notch-1-mediated gene regulation via p300.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenovirus E1A Proteins / metabolism
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Amino Acid Sequence
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Animals
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Basic Helix-Loop-Helix Transcription Factors
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Binding Sites
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Cell Line
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DNA-Binding Proteins / metabolism
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E1A-Associated p300 Protein
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Electrophoretic Mobility Shift Assay
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Green Fluorescent Proteins
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HeLa Cells
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Homeodomain Proteins / genetics
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Humans
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Immunoglobulin J Recombination Signal Sequence-Binding Protein
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Luminescent Proteins / genetics
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Luminescent Proteins / metabolism
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Mammals
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Mice
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Molecular Sequence Data
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Mutagenesis
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Nuclear Proteins / metabolism*
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Promoter Regions, Genetic
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Protein Binding
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Receptor, Notch1
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Receptors, Cell Surface*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Subcellular Fractions
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Trans-Activators / metabolism*
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Transcription Factor HES-1
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Transcription Factors*
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Transcription, Genetic
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Transcriptional Activation
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Tumor Suppressor Protein p53 / metabolism
Substances
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Adenovirus E1A Proteins
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Basic Helix-Loop-Helix Transcription Factors
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DNA-Binding Proteins
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Hes1 protein, mouse
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Homeodomain Proteins
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Immunoglobulin J Recombination Signal Sequence-Binding Protein
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Luminescent Proteins
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Membrane Proteins
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NOTCH1 protein, human
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Notch1 protein, mouse
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Nuclear Proteins
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RBPJ protein, human
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Rbpj protein, mouse
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Receptor, Notch1
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Receptors, Cell Surface
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Recombinant Fusion Proteins
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Trans-Activators
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Transcription Factor HES-1
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Transcription Factors
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Tumor Suppressor Protein p53
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Green Fluorescent Proteins
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HES1 protein, human
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E1A-Associated p300 Protein
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Ep300 protein, mouse