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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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Organization of fascin and vinculin in C2C12 cells adherent on adhesion receptor antibodies. (A) Cells were plated on surfaces coated with 50 μg/ml of the indicated anti–mouse antibodies for 1 h in serum-free medium and then fixed and stained for fascin or vinculin. (B) Fascin spike formation by primary mouse skeletal myoblasts on TSP-1. Wild-type or CD47- skeletal myoblasts were plated on 50 nM TSP-1 for 1 h and then stained for fascin. Both populations formed extensive fascin spikes. Bar: (A, panels 1–6) 15 μm; (A, vinculin-stained panel) 8 μm; (B) 10 μm.
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Figure 3: Organization of fascin and vinculin in C2C12 cells adherent on adhesion receptor antibodies. (A) Cells were plated on surfaces coated with 50 μg/ml of the indicated anti–mouse antibodies for 1 h in serum-free medium and then fixed and stained for fascin or vinculin. (B) Fascin spike formation by primary mouse skeletal myoblasts on TSP-1. Wild-type or CD47- skeletal myoblasts were plated on 50 nM TSP-1 for 1 h and then stained for fascin. Both populations formed extensive fascin spikes. Bar: (A, panels 1–6) 15 μm; (A, vinculin-stained panel) 8 μm; (B) 10 μm.

Mentions: In contrast, EDTA-disaggregated C2C12 cells spread fully on 281-2 antibody to syndecan-1 and displayed zones of F-actin ruffles and small projections at cell margins (Fig. 2). Cells trypsinized for 2 min, many of which retained cell surface syndecan-1 (Fig. 1 A), showed a less uniform response (Table and Fig. 3). Extensively trypsinized cells which lacked syndecan-1 attached at only background levels to the antibody and did not spread (data not shown). To establish that these responses were not due to differing affinities or avidities of individual antibody/antigen binding interactions, F-actin organization was examined on surfaces coated with different concentrations of the antibodies to β1 integrin, β3 integrin, CD47, or syndecan-1. The cells spread poorly on any antibody at 5 μg/ml coating concentration. Actin spikes and ruffles were organized by EDTA-dissociated cells on the 281-2 antibody in the concentration range 30–100 μg/ml and were not organized by cells adherent to the β1 or β3 antibodies even when they were coated at 100 μg/ml. Cells adherent on the CD47 antibody formed small F-actin ruffles on surfaces coated with ≥50 μg/ ml of the antibody (data not shown).


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

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

Organization of fascin and vinculin in C2C12 cells adherent on adhesion receptor antibodies. (A) Cells were plated on surfaces coated with 50 μg/ml of the indicated anti–mouse antibodies for 1 h in serum-free medium and then fixed and stained for fascin or vinculin. (B) Fascin spike formation by primary mouse skeletal myoblasts on TSP-1. Wild-type or CD47- skeletal myoblasts were plated on 50 nM TSP-1 for 1 h and then stained for fascin. Both populations formed extensive fascin spikes. Bar: (A, panels 1–6) 15 μm; (A, vinculin-stained panel) 8 μm; (B) 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199199&req=5

Figure 3: Organization of fascin and vinculin in C2C12 cells adherent on adhesion receptor antibodies. (A) Cells were plated on surfaces coated with 50 μg/ml of the indicated anti–mouse antibodies for 1 h in serum-free medium and then fixed and stained for fascin or vinculin. (B) Fascin spike formation by primary mouse skeletal myoblasts on TSP-1. Wild-type or CD47- skeletal myoblasts were plated on 50 nM TSP-1 for 1 h and then stained for fascin. Both populations formed extensive fascin spikes. Bar: (A, panels 1–6) 15 μm; (A, vinculin-stained panel) 8 μm; (B) 10 μm.
Mentions: In contrast, EDTA-disaggregated C2C12 cells spread fully on 281-2 antibody to syndecan-1 and displayed zones of F-actin ruffles and small projections at cell margins (Fig. 2). Cells trypsinized for 2 min, many of which retained cell surface syndecan-1 (Fig. 1 A), showed a less uniform response (Table and Fig. 3). Extensively trypsinized cells which lacked syndecan-1 attached at only background levels to the antibody and did not spread (data not shown). To establish that these responses were not due to differing affinities or avidities of individual antibody/antigen binding interactions, F-actin organization was examined on surfaces coated with different concentrations of the antibodies to β1 integrin, β3 integrin, CD47, or syndecan-1. The cells spread poorly on any antibody at 5 μg/ml coating concentration. Actin spikes and ruffles were organized by EDTA-dissociated cells on the 281-2 antibody in the concentration range 30–100 μg/ml and were not organized by cells adherent to the β1 or β3 antibodies even when they were coated at 100 μg/ml. Cells adherent on the CD47 antibody formed small F-actin ruffles on surfaces coated with ≥50 μg/ ml of the antibody (data not shown).

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