Abstract
We have investigated the role of protein kinase B (Akt) in the insulin-stimulated translocation of vesicles containing the insulin-responsive isoform of glucose transporter (GLUT4) to the plasma membrane of adipocytes. Previous reports have suggested that protein kinase B can bind to intracellular GLUT4 vesicles in an insulin-dependent manner, but the functional consequence of this translocation is not known. In this study we have artificially targeted constitutively active and kinase-inactive mutants of protein kinase B to intracellular GLUT4 vesicles by fusing them with the N-terminus of GLUT4 itself. We examined the effect of these mutants on the insulin-dependent translocation of the insulin-responsive amino peptidase IRAP (a bona fide GLUT4-vesicle-resident protein). A kinase-inactive protein kinase B targeted to GLUT4 vesicles was an extremely effective dominant-negative inhibitor of insulin-stimulated IRAP translocation to the plasma membrane. By contrast, a kinase-inactive protein kinase B expressed in the cytoplasm did not have an effect. The results suggest that protein kinase B has an important functional role at, or in the vicinity of, GLUT4 vesicles in the insulin-dependent translocation of those vesicles to the plasma membrane of adipocytes.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3T3 Cells
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Adipocytes / cytology
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Adipocytes / drug effects
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Adipocytes / metabolism*
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Aminopeptidases / drug effects
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Aminopeptidases / genetics
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Aminopeptidases / metabolism
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Animals
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Biomarkers
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Cell Compartmentation / drug effects
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Cell Compartmentation / genetics
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Cell Membrane / drug effects
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Cell Membrane / metabolism*
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Cell Membrane / ultrastructure
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Cystinyl Aminopeptidase
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DNA-Binding Proteins / drug effects
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Glucose / metabolism
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Glucose Transporter Type 4
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Insulin / metabolism*
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Insulin / pharmacology
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Mice
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Monosaccharide Transport Proteins / drug effects
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Monosaccharide Transport Proteins / genetics
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Monosaccharide Transport Proteins / metabolism*
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Muscle Proteins*
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Mutation / drug effects
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Mutation / genetics
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Protein Serine-Threonine Kinases*
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Protein Structure, Tertiary / drug effects
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Protein Structure, Tertiary / physiology
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Protein Transport / drug effects
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Protein Transport / physiology*
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Proto-Oncogene Proteins / drug effects
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism*
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Proto-Oncogene Proteins c-akt
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Recombinant Fusion Proteins / drug effects
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Transcription Factors / drug effects
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Transcription Factors / genetics
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Transcription Factors / metabolism
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Transport Vesicles / drug effects
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Transport Vesicles / metabolism*
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Transport Vesicles / ultrastructure
Substances
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Biomarkers
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DNA-Binding Proteins
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Glucose Transporter Type 4
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Insulin
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Monosaccharide Transport Proteins
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Muscle Proteins
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Proto-Oncogene Proteins
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Recombinant Fusion Proteins
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Slc2a4 protein, mouse
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Transcription Factors
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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Aminopeptidases
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Cystinyl Aminopeptidase
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leucyl-cystinyl aminopeptidase
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Glucose