Cell surface annexin II is a high affinity receptor for the alternatively spliced segment of tenascin-C.
We have investigated the binding of soluble tenascin-C (TN-C) to several cell lines using a radioligand binding assay. Specific binding was demonstrated to U-251MG human glioma cells and to a line of bovine aortic endothelial cells, but hamster fibroblasts showed no specific binding. Recombinant proteins corresponding to specific domains of ... TN-C were used to map the binding site(s) in TN-C. The alternatively spliced segment (TNfnA-D) inhibited the binding of native TN-C most strongly, and itself bound to glioma and endothelial cells. Scatchard analysis of TNfnA-D binding indicated 2-5 x 10(5) binding sites per cell, with an apparent 2 nM dissociation constant. The cell surface receptor for TNfnA-D was identified as a 35-kD protein on the basis of blot binding assays and affinity chromatography of membrane extracts on native TN-C and TNfnA-D columns. Protein sequencing indicated that this 35-kD receptor was annexin II. Annexin II is well characterized as a cytoplasmic protein, so it was surprising to find it as a presumably extracellular receptor for TN-C. To confirm that it was the 35-kD receptor, we obtained purified annexin II and demonstrated its binding to TNfnA-D and TN-C at nM concentrations. Antibodies to annexin II prominently stained the external surface of live endothelial cells and blocked the binding of TNfnA-D to the cells. Thus annexin II appears to be a receptor for the alternatively spliced segment of TN-C, and may mediate cellular responses to soluble TN-C in the extracellular matrix.
Mesh Terms:
Alternative Splicing, Amino Acid Sequence, Animals, Annexin A2, Binding Sites, Cattle, Cell Adhesion Molecules, Neuronal, Cells, Cultured, Endothelium, Extracellular Matrix Proteins, Glioma, Humans, Kinetics, Lung, Molecular Sequence Data, Nerve Tissue Proteins, Radioligand Assay, Receptors, Cell Surface, Recombinant Proteins, Sequence Analysis, Tenascin, Tumor Cells, Cultured
Alternative Splicing, Amino Acid Sequence, Animals, Annexin A2, Binding Sites, Cattle, Cell Adhesion Molecules, Neuronal, Cells, Cultured, Endothelium, Extracellular Matrix Proteins, Glioma, Humans, Kinetics, Lung, Molecular Sequence Data, Nerve Tissue Proteins, Radioligand Assay, Receptors, Cell Surface, Recombinant Proteins, Sequence Analysis, Tenascin, Tumor Cells, Cultured
J Cell Biol
Date: Jul. 01, 1994
PubMed ID: 7518469
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