Abstract
The intracellular localization of Shc proteins was analyzed by immunofluorescence and immunoelectron microscopy in normal cells and cells expressing the epidermal growth factor receptor or the EGFR/erbB2 chimera. In unstimulated cells, the immunolabeling was localized in the central perinuclear area of the cell and mostly associated with the cytosolic side of rough endoplasmic reticulum membranes. Upon epidermal growth factor treatment and receptor tyrosine kinase activation, the immunolabeling became peripheral and was found to be associated with the cytosolic surface of the plasma membrane and endocytic structures, such as coated pits and endosomes, and with the peripheral cytosol. Receptor activation in cells expressing phosphorylation-defective mutants of Shc and erbB-2 kinase showed that receptor autophosphorylation, but not Shc phosphorylation, is required for redistribution of Shc proteins. The rough endoplasmic reticulum localization of Shc proteins in unstimulated cells and their massive recruitment to the plasma membrane, endocytic structures, and peripheral cytosol following receptor tyrosine kinase activation could account for multiple putative functions of the adaptor protein.
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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Adaptor Proteins, Signal Transducing*
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Adaptor Proteins, Vesicular Transport*
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Animals
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Calcium-Calmodulin-Dependent Protein Kinases / metabolism
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Endoplasmic Reticulum / metabolism*
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Endoplasmic Reticulum / ultrastructure
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Enzyme Activation
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Epidermal Growth Factor / pharmacology
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ErbB Receptors / biosynthesis*
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ErbB Receptors / metabolism*
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Fluorescent Antibody Technique
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Mice
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Microscopy, Immunoelectron
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Phosphorylation
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Protein Biosynthesis
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Proteins / analysis
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Proteins / metabolism*
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Receptor Protein-Tyrosine Kinases / metabolism*
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Receptor, ErbB-2 / biosynthesis
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Receptor, ErbB-2 / metabolism
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Recombinant Fusion Proteins / biosynthesis
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Recombinant Fusion Proteins / metabolism
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Recombinant Proteins / analysis
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Recombinant Proteins / biosynthesis
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Recombinant Proteins / metabolism
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Shc Signaling Adaptor Proteins
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Src Homology 2 Domain-Containing, Transforming Protein 1
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Subcellular Fractions / metabolism
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Subcellular Fractions / ultrastructure
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Transfection
Substances
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Adaptor Proteins, Signal Transducing
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Adaptor Proteins, Vesicular Transport
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Proteins
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Recombinant Fusion Proteins
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Recombinant Proteins
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Shc Signaling Adaptor Proteins
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Shc1 protein, mouse
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Src Homology 2 Domain-Containing, Transforming Protein 1
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Epidermal Growth Factor
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ErbB Receptors
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Receptor Protein-Tyrosine Kinases
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Receptor, ErbB-2
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Calcium-Calmodulin-Dependent Protein Kinases