Abstract
We have previously identified angiomotin by its ability to bind to and mediate the anti-angiogenic properties of angiostatin. In vivo and in vitro data indicate an essential role of angiomotin in endothelial cell motility. Here we show that angiostatin binds angiomotin on the cell surface and provide evidence for a transmembrane model for the topology of both p80 and p130 angiomotin isoforms. Immunofluorescence analysis shows that angiomotin co-localized with ZO-1 in cell-cell contacts in endothelial cells in vitro and in angiogenic blood vessels of the postnatal mouse retina in vivo. Transfection of p80 as well as p130 angiomotin in Chinese hamster ovary cells resulted in junctional localization of both isoforms. Furthermore, p130 angiomotin could recruit ZO-1 to actin stress fibers. The p130 but not p80 isoform could be coprecipitated with MAGI-1b, a component of endothelial tight junctions. Paracellular permeability, as measured by diffusion of fluorescein isothiocyanate-dextran, was reduced by p80 and p130 angiomotin expression with 70 and 88%, respectively, compared with control. Angiostatin did not have any effect on cell permeability but inhibited the migration of angiomotin-expressing cells in the Boyden chamber assay. We conclude that angiomotin, in addition to controlling cell motility, may play a role in the assembly of endothelial cell-cell junctions.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Alternative Splicing
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Angiomotins
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Angiopoietin-Like Protein 1
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Angiostatins / chemistry
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Angiostatins / metabolism
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Animals
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Biotinylation
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Blotting, Western
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CHO Cells
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Capillaries / cytology
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Carrier Proteins / physiology*
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Cattle
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Cell Communication
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Cell Membrane / metabolism
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Cell Movement
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Computational Biology
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Cricetinae
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Cross-Linking Reagents / pharmacology
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Diffusion
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Endothelial Cells / cytology*
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Endothelium, Vascular / cytology
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Fluorescein-5-isothiocyanate / pharmacology
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Gene Expression Regulation*
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HeLa Cells
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Humans
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Immunoprecipitation
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Intercellular Signaling Peptides and Proteins / physiology*
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Intracellular Signaling Peptides and Proteins
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Membrane Proteins / chemistry
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Membrane Proteins / metabolism
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Mice
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Microfilament Proteins
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Microscopy, Fluorescence
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Octoxynol
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Permeability
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Phosphoproteins / metabolism
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Plasmids / metabolism
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Polyethylene Glycols / pharmacology
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Protein Binding
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Protein Isoforms
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Protein Structure, Tertiary
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Proteins / metabolism
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Retina / metabolism
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Retinoblastoma-Like Protein p130 / metabolism
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Time Factors
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Trypsin / pharmacology
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Zonula Occludens-1 Protein
Substances
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AMOT protein, human
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AMOTL1 protein, human
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Amotl1 protein, mouse
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Angiomotins
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Angiopoietin-Like Protein 1
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Carrier Proteins
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Cross-Linking Reagents
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Intercellular Signaling Peptides and Proteins
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Intracellular Signaling Peptides and Proteins
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Membrane Proteins
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Microfilament Proteins
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Phosphoproteins
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Protein Isoforms
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Proteins
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Rbl2 protein, mouse
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Retinoblastoma-Like Protein p130
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TJP1 protein, human
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Tjp1 protein, mouse
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WDR48 protein, human
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Zonula Occludens-1 Protein
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Polyethylene Glycols
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Angiostatins
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Octoxynol
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Nonidet P-40
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Trypsin
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Fluorescein-5-isothiocyanate