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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 F-actin and fascin in cells adherent on TSP-1 or adhesion receptor antibodies. (A) EDTA-released mouse embryonic fibroblast cells (MEF) were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of the indicated antibodies as used in Fig. 1 and Fig. 2 for 1 h in serum-free medium and then fixed and stained for fascin. (B) EDTA-released HLMECs were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of antibodies to human adhesion receptors for 1 h in serum-free medium and then fixed and stained for F-actin or fascin. Bars, 20 μm.
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Figure 4: Organization of F-actin and fascin in cells adherent on TSP-1 or adhesion receptor antibodies. (A) EDTA-released mouse embryonic fibroblast cells (MEF) were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of the indicated antibodies as used in Fig. 1 and Fig. 2 for 1 h in serum-free medium and then fixed and stained for fascin. (B) EDTA-released HLMECs were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of antibodies to human adhesion receptors for 1 h in serum-free medium and then fixed and stained for F-actin or fascin. Bars, 20 μm.

Mentions: We extended the analysis of receptor ligation by antibodies to additional cell types which show different degrees of spreading on TSP-1 substrata. MEF-1 mouse fibroblasts spread and form fascin spikes and filopodia on TSP-1 (Fig. 4 A). Cells remained rounded with diffuse fascin when plated on the anti–mouse β1, β3, or CD47 antibodies (shown for β1 only, Fig. 4 A). When plated on the 281-2 antibody to mouse syndecan-1, MEF-1 formed broad circumferential lamellae containing radial F-actin ribs which corresponded to sites where fascin and actin were bundled (Fig. 4 A). HLMECs express fascin and syndecan-1 but as reported for endothelial cells from other blood vessel sources, do not spread on TSP-1 (Fig. 4 B; Lawler et al. 1988; Murphy-Ullrich and Hook 1989; Mertens et al. 1992; Adams, J.C., unpublished data). The cells show specific enrichment of F-actin and fascin structures at cell margins when attached to TSP-1 (Fig. 4 B). HLMECs attached to surfaces coated with antibodies to human β1, β3, CD47, or syndecan-1 and showed most microfilament organization on the β1 antibody. However, fascin was completely diffuse in these cells and was not enriched at cell margins (Fig. 4 B; data not shown). On either of two antibodies to human syndecan-1, HLMECs formed short marginal structures which contained F-actin and fascin (shown for BB4 antibody only, Fig. 4 B). We also examined fascin localization in HLMECs plated on surfaces coated with SMO antibody to human CD36 (Hogg et al. 1984). The cells remained rounded and smooth edged and did not show specific localization of fascin (data not shown). Neither the BRIC 126, B6H12, nor 2E11 antibodies to human CD47 caused fascin–actin bundling (Fig. 4 B; data not shown). Thus, in several cell types from different tissue sources, ligation of syndecan-1 specifically promoted the organization of cortical fascin independently of the degree of cell spreading.


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 F-actin and fascin in cells adherent on TSP-1 or adhesion receptor antibodies. (A) EDTA-released mouse embryonic fibroblast cells (MEF) were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of the indicated antibodies as used in Fig. 1 and Fig. 2 for 1 h in serum-free medium and then fixed and stained for fascin. (B) EDTA-released HLMECs were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of antibodies to human adhesion receptors for 1 h in serum-free medium and then fixed and stained for F-actin or fascin. Bars, 20 μm.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 4: Organization of F-actin and fascin in cells adherent on TSP-1 or adhesion receptor antibodies. (A) EDTA-released mouse embryonic fibroblast cells (MEF) were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of the indicated antibodies as used in Fig. 1 and Fig. 2 for 1 h in serum-free medium and then fixed and stained for fascin. (B) EDTA-released HLMECs were plated on 50 nM TSP-1 or surfaces coated with 50 μg/ ml of antibodies to human adhesion receptors for 1 h in serum-free medium and then fixed and stained for F-actin or fascin. Bars, 20 μm.
Mentions: We extended the analysis of receptor ligation by antibodies to additional cell types which show different degrees of spreading on TSP-1 substrata. MEF-1 mouse fibroblasts spread and form fascin spikes and filopodia on TSP-1 (Fig. 4 A). Cells remained rounded with diffuse fascin when plated on the anti–mouse β1, β3, or CD47 antibodies (shown for β1 only, Fig. 4 A). When plated on the 281-2 antibody to mouse syndecan-1, MEF-1 formed broad circumferential lamellae containing radial F-actin ribs which corresponded to sites where fascin and actin were bundled (Fig. 4 A). HLMECs express fascin and syndecan-1 but as reported for endothelial cells from other blood vessel sources, do not spread on TSP-1 (Fig. 4 B; Lawler et al. 1988; Murphy-Ullrich and Hook 1989; Mertens et al. 1992; Adams, J.C., unpublished data). The cells show specific enrichment of F-actin and fascin structures at cell margins when attached to TSP-1 (Fig. 4 B). HLMECs attached to surfaces coated with antibodies to human β1, β3, CD47, or syndecan-1 and showed most microfilament organization on the β1 antibody. However, fascin was completely diffuse in these cells and was not enriched at cell margins (Fig. 4 B; data not shown). On either of two antibodies to human syndecan-1, HLMECs formed short marginal structures which contained F-actin and fascin (shown for BB4 antibody only, Fig. 4 B). We also examined fascin localization in HLMECs plated on surfaces coated with SMO antibody to human CD36 (Hogg et al. 1984). The cells remained rounded and smooth edged and did not show specific localization of fascin (data not shown). Neither the BRIC 126, B6H12, nor 2E11 antibodies to human CD47 caused fascin–actin bundling (Fig. 4 B; data not shown). Thus, in several cell types from different tissue sources, ligation of syndecan-1 specifically promoted the organization of cortical fascin independently of the degree of cell spreading.

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