Abstract
Vinexin is a recently identified cytoskeletal protein and plays a key role in the regulation of cytoskeletal organization and signal transduction. Vinexin localizes at sites of cell-extracellular matrix adhesion in NIH3T3 fibroblasts and at sites of cell-cell contact in epithelial LLC-PK1 cells. Expression of vinexin promotes the formation of actin stress fiber, but the role of vinexin at sites of cell-cell contact is unclear. Here we identified lp-dlg/KIAA0583 as a novel binding partner for vinexin by using yeast two-hybrid screening. lp-dlg/KIAA0583 has a NH2-terminal coiled-coil-like domain, in addition to four PDZ domains, an Src homology (SH) 3 domain, and a guanylate kinase domain, which are conserved structures in membrane-associated guanylate kinase family proteins. The third SH3 domain of vinexin bound to the region between the second and third PDZ domain of lp-dlg, which contains a proline-rich sequence. lp-dlg colocalized with vinexin at sites of cell-cell contact in LLC-PK1 cells. Furthermore, lp-dlg colocalized with beta-catenin, a major adherens junction protein, in LLC-PK1 cells. Co-immunoprecipitation experiments revealed that both endogenous and epitope-tagged deletion mutants of lp-dlg/KIAA0583 associated with beta-catenin. We also showed that these three proteins could form a ternary complex. Together these findings suggest that lp-dlg/KIAA0583 is a novel scaffolding protein that can link the vinexin-vinculin complex and beta-catenin at sites of cell-cell contact.
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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Amino Acid Sequence
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Animals
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Blotting, Northern
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COS Cells
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Cell Communication
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Cell Line
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Cell Membrane / metabolism
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Cloning, Molecular
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Cytoskeletal Proteins / chemistry
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Cytoskeletal Proteins / metabolism*
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DNA, Complementary / metabolism
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Discs Large Homolog 1 Protein
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Epithelial Cells / metabolism
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Gene Deletion
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Genes, Tumor Suppressor
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Glutathione Transferase / metabolism
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Guanylate Kinases
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Humans
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Membrane Proteins / chemistry*
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Membrane Proteins / metabolism*
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Mice
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Models, Genetic
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Molecular Sequence Data
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Muscle Proteins / chemistry
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Muscle Proteins / metabolism*
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Nucleoside-Phosphate Kinase / chemistry*
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Nucleoside-Phosphate Kinase / genetics
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Polymerase Chain Reaction
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Precipitin Tests
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Protein Binding
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Protein Structure, Tertiary
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Proteins / genetics*
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Recombinant Fusion Proteins / metabolism
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Tissue Distribution
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Trans-Activators / chemistry
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Trans-Activators / metabolism*
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Tumor Suppressor Proteins*
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Two-Hybrid System Techniques
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beta Catenin
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src Homology Domains
Substances
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Adaptor Proteins, Signal Transducing
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CTNNB1 protein, human
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CTNNB1 protein, mouse
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Cytoskeletal Proteins
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DLG1 protein, human
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DLG5 protein, human
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DNA, Complementary
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Discs Large Homolog 1 Protein
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Dlg1 protein, mouse
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Membrane Proteins
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Muscle Proteins
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Proteins
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Recombinant Fusion Proteins
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SORBS3 protein, human
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Sorbs3 protein, mouse
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Trans-Activators
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Tumor Suppressor Proteins
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beta Catenin
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Glutathione Transferase
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Nucleoside-Phosphate Kinase
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Guanylate Kinases