Centrosomal P4.1-associated protein is a new member of transcriptional coactivators for nuclear factor-kappaB.
Nuclear factor-kappaB (NF-kappaB) is a transcription factor important for various cellular events such as inflammation, immune response, proliferation, and apoptosis. In this study, we performed a yeast two-hybrid screening using the N-terminal domain of the p65 subunit (RelA) of NF-kappaB as bait and isolated centrosomal P4.1-associated protein (CPAP) as a ... candidate for a RelA-associating partner. Glutathione S-transferase pull-down assays and co-immunoprecipitation experiments followed by Western blotting also showed association of CPAP with RelA. When overexpressed, CPAP enhanced NF-kappaB-dependent transcription induced by tumor necrosis factor-alpha (TNFalpha). Reduction of the protein level of endogenous CPAP by RNA interference resulted in decreased activation of NF-kappaB by TNFalpha. After treatment with TNFalpha, a portion of CPAP was observed to accumulate in the nucleus, although CPAP was found primarily in the cytoplasm without any stimulation. Moreover, CPAP was observed in a complex recruited to the transcriptional promoter region containing the NF-kappaB-binding motif. One hybrid assay showed that CPAP has the potential to activate gene expression when tethered to the transcriptional promoter. These data suggest that CPAP functions as a coactivator of NF-kappaB-mediated transcription. Since a physiological interaction between CPAP and the coactivator p300/CREB-binding protein was also observed and synergistic activation of NF-kappaB-mediated transcription was achieved by these proteins, CPAP-dependent transcriptional activation is likely to include p300/CREB-binding protein.
Mesh Terms:
Amino Acid Motifs, Binding Sites, Blotting, Western, Cell Line, Cell Line, Tumor, Cell Nucleus, Centrosome, Cytoplasm, DNA, Fluorescent Antibody Technique, Indirect, Gene Expression Regulation, Genes, Reporter, Glutathione Transferase, Humans, Immunoblotting, Immunoprecipitation, Luciferases, Microtubule-Associated Proteins, Models, Genetic, NF-kappa B, Plasmids, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, RNA Interference, Recombinant Proteins, Reverse Transcriptase Polymerase Chain Reaction, Time Factors, Transcription Factor RelA, Transcription, Genetic, Transcriptional Activation, Transfection, Tumor Necrosis Factor-alpha, Two-Hybrid System Techniques
Amino Acid Motifs, Binding Sites, Blotting, Western, Cell Line, Cell Line, Tumor, Cell Nucleus, Centrosome, Cytoplasm, DNA, Fluorescent Antibody Technique, Indirect, Gene Expression Regulation, Genes, Reporter, Glutathione Transferase, Humans, Immunoblotting, Immunoprecipitation, Luciferases, Microtubule-Associated Proteins, Models, Genetic, NF-kappa B, Plasmids, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, RNA Interference, Recombinant Proteins, Reverse Transcriptase Polymerase Chain Reaction, Time Factors, Transcription Factor RelA, Transcription, Genetic, Transcriptional Activation, Transfection, Tumor Necrosis Factor-alpha, Two-Hybrid System Techniques
J. Biol. Chem.
Date: Apr. 01, 2005
PubMed ID: 15687488
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