Abstract
TAK1 mitogen-activated protein kinase kinase kinase (MAP3K) is activated by its specific activator, TAK1-binding protein 1 (TAB1). A constitutively active TAK1 mutant has not yet been generated due to the indispensable requirement of TAB1 for TAK1 kinase activity. In this study, we generated a novel constitutively active TAK1 by fusing its kinase domain to the minimal TAK1-activation domain of TAB1. Co-immunoprecipitation assay demonstrated that these domains interacted intra-molecularly. The TAK1-TAB1 fusion protein showed a significant MAP3K activity in vitro and activated c-Jun N-terminal kinase/p38 MAPKs and IkappaB kinase in vivo, which was followed by increased production of interleukin-6. These results indicate that the fusion protein is useful for characterizing the physiological roles of the TAK1-TAB1 complex.
MeSH terms
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Adaptor Proteins, Signal Transducing*
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Binding Sites
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Blotting, Western
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Carrier Proteins / chemistry*
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Carrier Proteins / metabolism
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DNA, Complementary / metabolism
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Enzyme-Linked Immunosorbent Assay
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HeLa Cells
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Humans
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Interleukin-6 / metabolism
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Interleukin-8 / metabolism
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Intracellular Signaling Peptides and Proteins*
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JNK Mitogen-Activated Protein Kinases
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MAP Kinase Kinase Kinases / chemistry*
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MAP Kinase Kinase Kinases / metabolism
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MAP Kinase Signaling System
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Mitogen-Activated Protein Kinases / metabolism
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Models, Genetic
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Mutation
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NF-kappa B / metabolism*
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Precipitin Tests
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Protein Binding
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Protein Structure, Tertiary
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Recombinant Fusion Proteins / metabolism
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Transcription Factor AP-1 / metabolism*
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Transfection
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p38 Mitogen-Activated Protein Kinases
Substances
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Adaptor Proteins, Signal Transducing
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Carrier Proteins
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DNA, Complementary
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Interleukin-6
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Interleukin-8
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Intracellular Signaling Peptides and Proteins
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NF-kappa B
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Recombinant Fusion Proteins
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TAB1 protein, MAPKKK activator, vertebrate
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TAB1 protein, human
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Transcription Factor AP-1
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JNK Mitogen-Activated Protein Kinases
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Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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MAP Kinase Kinase Kinases
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MAP kinase kinase kinase 7