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A role for syndecan-1 in coupling fascin spike formation by thrombospondin-1.

Adams JC, Kureishy N, Taylor AL - J. Cell Biol. (2001)

Bottom Line: The underlying molecular mechanism depends on glycosaminoglycan (GAG) modification of the syndecan-1 core protein at residues S45 or S47 for cell membrane spreading and on the VC2 region of the cytoplasmic domain for spreading and fascin spike formation.Expression of the VC2 deletion mutant or GAG-negative syndecan-1 showed that syndecan-1 is necessary in spreading and fascin spike formation by C2C12 cells on TSP-1.These results establish a novel role for syndecan-1 protein in coupling a physiological matrix ligand to formation of a specific matrix contact structure.

View Article: PubMed Central - PubMed

Affiliation: Medical Research Council Laboratory for Molecular Cell Biology and Department of Biochemistry and Molecular Biology, University College London, London WC1E 6BT, United Kingdom. dmcbjca@ucl.ac.uk

ABSTRACT
An important role of cell matrix adhesion receptors is to mediate transmembrane coupling between extracellular matrix attachment, actin reorganization, and cell spreading. Thrombospondin (TSP)-1 is a modulatory component of matrix expressed during development, immune response, or wound repair. Cell adhesion to TSP-1 involves formation of biochemically distinct matrix contacts based on stable fascin spikes. The cell surface adhesion receptors required have not been identified. We report here that antibody clustering of syndecan-1 proteoglycan specifically transduces organization of cortical actin and fascin bundles in several cell types. Transfection of COS-7 cells with syndecan-1 is sufficient to stimulate cell spreading, fascin spike assembly, and extensive protrusive lateral ruffling on TSP-1 or on syndecan-1 antibody. The underlying molecular mechanism depends on glycosaminoglycan (GAG) modification of the syndecan-1 core protein at residues S45 or S47 for cell membrane spreading and on the VC2 region of the cytoplasmic domain for spreading and fascin spike formation. Expression of the VC2 deletion mutant or GAG-negative syndecan-1 showed that syndecan-1 is necessary in spreading and fascin spike formation by C2C12 cells on TSP-1. These results establish a novel role for syndecan-1 protein in coupling a physiological matrix ligand to formation of a specific matrix contact structure.

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Expression of syndecan-1 specifically promotes cell spreading and formation of fascin spikes in TSP-1–adherent COS-7 cells. COS-7 cells were transfected with syndecan-1 and EGFP-fascin expression plasmids for 48 h. EDTA-released cells were plated on 50 nM TSP-1, fibro nectin, or 40 μg/ml anti–mouse syndecan-1 antibody in serum-free medium for 1 h and then fixed and stained for F-actin. Results shown are representative of three independent experiments, and ≥100 transfected cells were scored per experiment. The morphological features used for quantitation are also indicated. These were cell area measured by spread cell edge, and numbers and lengths of the radial fascin–actin bundles that form spikes and ribs. Examples of these bundles are marked with asteriks in two of the panels. Bars: (top two panels) 16 μm; (bottom six panels) 5 μm.
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Figure 5: Expression of syndecan-1 specifically promotes cell spreading and formation of fascin spikes in TSP-1–adherent COS-7 cells. COS-7 cells were transfected with syndecan-1 and EGFP-fascin expression plasmids for 48 h. EDTA-released cells were plated on 50 nM TSP-1, fibro nectin, or 40 μg/ml anti–mouse syndecan-1 antibody in serum-free medium for 1 h and then fixed and stained for F-actin. Results shown are representative of three independent experiments, and ≥100 transfected cells were scored per experiment. The morphological features used for quantitation are also indicated. These were cell area measured by spread cell edge, and numbers and lengths of the radial fascin–actin bundles that form spikes and ribs. Examples of these bundles are marked with asteriks in two of the panels. Bars: (top two panels) 16 μm; (bottom six panels) 5 μm.

Mentions: The vector-transfected cells attached to TSP-1 remained rounded, and showed concentrations of actin and fascin at cell margins indistinguishable from those of untransfected cells (Fig. 5). In marked contrast, syndecan-1 transfectants showed greatly enhanced spreading on TSP-1. The cells spread to form adherent lamellipodial sheets with marginal F-actin and fascin-containing projections of varying lengths (Fig. 5). The syndecan-1 transfectants attached on the syndecan-1 antibody also showed greatly increased spreading which involved the formation of flatly spread lamellae in which radial projections containing F-actin and fascin were present at spread margins or extended as individual projections beyond the edge of the lamellae (Fig. 5). This effect involved a dramatic increase in the number of fascin spikes per cell (see Fig. 7 A). Cell membrane spreading involved a threefold increase in cell area (statistically significant at P = 0.0001; see Fig. 7 A). To examine the specificity of this effect, the syndecan-1 transfectants were plated on another matrix ligand, fibronectin. No increase in cell area or alteration in major F-actin structures was observed (Fig. 5; data not shown). Thus, the effects of syndecan-1 depend on its ligation by specific ligands and are not simply a consequence of the surface expression of syndecan-1.


A role for syndecan-1 in coupling fascin spike formation by thrombospondin-1.

Adams JC, Kureishy N, Taylor AL - J. Cell Biol. (2001)

Expression of syndecan-1 specifically promotes cell spreading and formation of fascin spikes in TSP-1–adherent COS-7 cells. COS-7 cells were transfected with syndecan-1 and EGFP-fascin expression plasmids for 48 h. EDTA-released cells were plated on 50 nM TSP-1, fibro nectin, or 40 μg/ml anti–mouse syndecan-1 antibody in serum-free medium for 1 h and then fixed and stained for F-actin. Results shown are representative of three independent experiments, and ≥100 transfected cells were scored per experiment. The morphological features used for quantitation are also indicated. These were cell area measured by spread cell edge, and numbers and lengths of the radial fascin–actin bundles that form spikes and ribs. Examples of these bundles are marked with asteriks in two of the panels. Bars: (top two panels) 16 μm; (bottom six panels) 5 μm.
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Related In: Results  -  Collection

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Figure 5: Expression of syndecan-1 specifically promotes cell spreading and formation of fascin spikes in TSP-1–adherent COS-7 cells. COS-7 cells were transfected with syndecan-1 and EGFP-fascin expression plasmids for 48 h. EDTA-released cells were plated on 50 nM TSP-1, fibro nectin, or 40 μg/ml anti–mouse syndecan-1 antibody in serum-free medium for 1 h and then fixed and stained for F-actin. Results shown are representative of three independent experiments, and ≥100 transfected cells were scored per experiment. The morphological features used for quantitation are also indicated. These were cell area measured by spread cell edge, and numbers and lengths of the radial fascin–actin bundles that form spikes and ribs. Examples of these bundles are marked with asteriks in two of the panels. Bars: (top two panels) 16 μm; (bottom six panels) 5 μm.
Mentions: The vector-transfected cells attached to TSP-1 remained rounded, and showed concentrations of actin and fascin at cell margins indistinguishable from those of untransfected cells (Fig. 5). In marked contrast, syndecan-1 transfectants showed greatly enhanced spreading on TSP-1. The cells spread to form adherent lamellipodial sheets with marginal F-actin and fascin-containing projections of varying lengths (Fig. 5). The syndecan-1 transfectants attached on the syndecan-1 antibody also showed greatly increased spreading which involved the formation of flatly spread lamellae in which radial projections containing F-actin and fascin were present at spread margins or extended as individual projections beyond the edge of the lamellae (Fig. 5). This effect involved a dramatic increase in the number of fascin spikes per cell (see Fig. 7 A). Cell membrane spreading involved a threefold increase in cell area (statistically significant at P = 0.0001; see Fig. 7 A). To examine the specificity of this effect, the syndecan-1 transfectants were plated on another matrix ligand, fibronectin. No increase in cell area or alteration in major F-actin structures was observed (Fig. 5; data not shown). Thus, the effects of syndecan-1 depend on its ligation by specific ligands and are not simply a consequence of the surface expression of syndecan-1.

Bottom Line: The underlying molecular mechanism depends on glycosaminoglycan (GAG) modification of the syndecan-1 core protein at residues S45 or S47 for cell membrane spreading and on the VC2 region of the cytoplasmic domain for spreading and fascin spike formation.Expression of the VC2 deletion mutant or GAG-negative syndecan-1 showed that syndecan-1 is necessary in spreading and fascin spike formation by C2C12 cells on TSP-1.These results establish a novel role for syndecan-1 protein in coupling a physiological matrix ligand to formation of a specific matrix contact structure.

View Article: PubMed Central - PubMed

Affiliation: Medical Research Council Laboratory for Molecular Cell Biology and Department of Biochemistry and Molecular Biology, University College London, London WC1E 6BT, United Kingdom. dmcbjca@ucl.ac.uk

ABSTRACT
An important role of cell matrix adhesion receptors is to mediate transmembrane coupling between extracellular matrix attachment, actin reorganization, and cell spreading. Thrombospondin (TSP)-1 is a modulatory component of matrix expressed during development, immune response, or wound repair. Cell adhesion to TSP-1 involves formation of biochemically distinct matrix contacts based on stable fascin spikes. The cell surface adhesion receptors required have not been identified. We report here that antibody clustering of syndecan-1 proteoglycan specifically transduces organization of cortical actin and fascin bundles in several cell types. Transfection of COS-7 cells with syndecan-1 is sufficient to stimulate cell spreading, fascin spike assembly, and extensive protrusive lateral ruffling on TSP-1 or on syndecan-1 antibody. The underlying molecular mechanism depends on glycosaminoglycan (GAG) modification of the syndecan-1 core protein at residues S45 or S47 for cell membrane spreading and on the VC2 region of the cytoplasmic domain for spreading and fascin spike formation. Expression of the VC2 deletion mutant or GAG-negative syndecan-1 showed that syndecan-1 is necessary in spreading and fascin spike formation by C2C12 cells on TSP-1. These results establish a novel role for syndecan-1 protein in coupling a physiological matrix ligand to formation of a specific matrix contact structure.

Show MeSH
Related in: MedlinePlus