COMMUNICATION:
Syndecan-4 Proteoglycan Regulates the Distribution and Activity of Protein Kinase C*

(Received for publication, January 24, 1997)

Eok-Soo Oh , Anne Woods Dagger and John R. Couchman

From the Department of Cell Biology, University of Alabama at Birmingham, Birmingham, Alabama 35294

ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
FOOTNOTES
ACKNOWLEDGEMENTS
REFERENCES


ABSTRACT

During cell-matrix adhesion, both tyrosine and serine/threonine kinases are activated. Integrin ligation correlates with tyrosine phosphorylation, whereas the later stages of spreading and focal adhesion and stress fiber formation in primary fibroblasts requires interactions of cell surface proteoglycan with heparin-binding moieties. This correlates with protein kinase C (PKC) activation, and PKCalpha can become localized to focal adhesions in normal, but not transformed, cells. PKC activation has been thought to be downstream of initial receptor-ligand interactions. We now show, however, that syndecan-4 transmembrane heparan sulfate proteoglycan and PKC co-immunoprecipitate and co-patch in vivo. The core protein of syndecan-4 can directly bind the catalytic domain of PKCalpha and potentiate its activation by phospholipid mediators. It can also directly activate PKCalpha in the absence of other mediators. This activity resides in the sequence LGKKPIYKK in the center of the short cytoplasmic domain, and other syndecans lack this sequence and PKC regulatory properties. Syndecan-4 is a focal adhesion component, and this interaction may both localize PKC and amplify its activity at sites of forming adhesions. This represents the first report of direct transmembrane signaling through cell surface proteoglycans.


INTRODUCTION

In cell-matrix interactions, as with many cellular responses to ligands, both tyrosine and serine/threonine kinases participate. During adhesion to fibronectin, integrin clustering activates tyrosine kinases (1-3), but we, and others, have noted that full spreading (4) and the formation of stress fibers and focal adhesions requires additional signals (3, 5-12). Adhesion to the "cell-binding" 105-kDa fragment of fibronectin through ligation of integrin alpha 5beta 1 is sufficient only for attachment and spreading in anchorage-dependent primary fibroblasts in the absence of serum and protein synthesis (8-11). To drive the cytoskeletal and receptor clustering that accompany the formation of stress fibers and focal adhesions, an additional activation of protein kinase C (PKC)1 is needed (2, 8, 9). This can be provided by addition of heparin-binding fibronectin moieties (8-11), either of the whole heparin-binding domain of fibronectin or of a peptide PRARI from this domain. These agents appear to signal through a cell surface heparan sulfate proteoglycan, since treatment with heparinase prevents the response (11), and mutant cells lacking (13), or having undersulfated cell surface heparan sulfate proteoglycans (14), have reduced capacity to form focal adhesions or stress fibers. Similarly, addition of PRARI peptides to synovial fibroblasts prespread on substrates of 105-kDa fragment markedly increased the size of vinculin-positive adhesion plaques (12). The need for addition of heparin-binding agents can be circumvented by treatment of cells with active, but not inactive, phorbol esters (8). In addition, there is downstream activation of the RhoA G-protein (15), which may initiate a contractile response (3).

Syndecan-4, one of the four members of transmembrane matrix-binding proteoglycans (16-18), becomes inserted into the focal adhesions of a range of cell types (19) when PKC is activated (20). PKCalpha is also localized in focal adhesions of normal cells (21) and is presumably anchored there, but the mechanism of anchorage has not been elucidated. Although activation of PKC is usually considered a downstream effect following receptor-ligand interactions, these previous studies led us to investigate whether syndecan-4 core protein could directly bind PKC and effect its localization to forming adhesions. We now report the first example of a mechanism for signaling through a transmembrane proteoglycan. The cytoplasmic domain of syndecan-4 core protein can directly regulate both the localization and activity of PKC through interactions of a sequence unique to syndecan-4 with the catalytic domain of PKC.


EXPERIMENTAL PROCEDURES

Binding Assays

For co-immunoprecipitation experiments, subconfluent rat embryo fibroblasts (REF) were scraped (approx 107 cells/immunoprecipitate) into 10 ml of lysis buffer (1% Triton X-100 in 50 mM Tris-HCl, pH 7.5, containing 50 mM NaCl, 0.2 mM phenylmethylsulfonyl fluoride, 1 mM benzamidine HCl, 1 mM leupeptin, 750 µM CaCl2), precleared by centrifugation, and incubated for 30 min at 4 °C with protein A-Sepharose beads (Pharmacia Biotech Inc.) pre-blocked (30 min) with 10% fetal calf serum in phosphate-buffered saline. After centrifugation, supernatants were sequentially incubated (30 min each) with 20 µg/ml monoclonal anti-syndecan-4 150.9 IgG raised against its NH2 terminus, 20 µg/ml rabbit anti-mouse IgG (Dako Corp. Carpinteria, CA) and fresh preblocked protein A-Sepharose beads. Beads were centrifuged, washed eight times with lysis buffer, boiled for 3 min, and released material immunoblotted with pan-specific anti-PKC recognizing the COOH terminus of the catalytic domains of PKCalpha , PKCbeta I, PKCbeta II, PKCgamma , and PKCdelta (Upstate Biotechnology, Inc., Lake Placid, NY) following 3-15% SDS-PAGE. Bound antibody was detected by goat anti-rabbit alkaline phosphatase (Bio-Rad). Some cultures were pretreated before lysis with phorbol 12-myristate 13-acetate (PMA: 200 nM, 15 min); in others 4V or scrambled peptide (50 µg/ml) was added at lysis and was present throughout. To demonstrate co-patching in vivo, REF were plated for 20 min onto coverslips coated with fibronectin (19), treated sequentially with 150.9 IgG and goat anti-mouse fluorescein isothiocyanate (Organon Teknika Corp., Durham, NC), warmed for 20 min in the presence or absence of 200 nM PMA (22), and stained with anti-PKC followed by goat anti-rabbit conjugated to Texas Red (Organon Teknika) after fixation and permeabilization (19). No breakthrough or inappropriate cross-reactivity of antibodies was detected in controls. For affinity chromatography on Affi-Gel-102 to which peptides were coupled through NH2-terminal cysteine by sulfo-LC-SPDP (Pierce), REF (approx 5 × 106 cells/sample) were scraped into 6 ml of lysis buffer lacking calcium, centrifuged (100,000 × g, 1 h), and CaCl2 (750 µM) and PMA (200 nM) were added to the supernatant as indicated. Columns were washed with 50 mM NaCl and eluted with 1 mM EDTA, or 1 M NaCl, released material being concentrated with 10% trichloroacetic acid, and immunoblotted with anti-PKC following 7.5% SDS-PAGE. Bound antibody was detected by chemiluminescence (ECL; Amersham Corp.). In solid phase binding assays (Pierce), enzyme-linked immunosorbent assay plate wells were coated with peptides (1 µg/well in NaHCO3, pH 9.6, 4 °C, overnight), washed, and unoccupied sites blocked with 5% milk (1 h). Second, PKCalpha (10 ng) was allowed to bind (37 °C, 30 min) in the presence or absence (for binding conditions, see legend to Fig. 1D) of phospholipid (50 µg/ml PS, 5 µg/ml DL) and/or 1 mM CaCl2. Third, bound enzyme was activated with 0.2 mg/ml PS, 0.02 mg/ml DL, 750 µM CaCl2 to phosphorylate added dye-labeled serine-substituted PKC pseudosubstrate (16 ng/well, 37 °C for 30 min). Phosphorylation was monitored by absorbance at 570 nm following affinity separation. Absorbance is normally proportional to the amount of PKCalpha bound.


Fig. 1. Syndecan-4 binds PKC. A, immunoblotting of syndecan-4 immunoprecipitates with anti-PKC shows co-precipitation in the presence (lane 2), but not absence (lane 1), of PMA. This was reduced by 4V (lane 5) but not scrambled peptide (lane 6). No binding occurred to corresponding noncoated protein A beads (lanes 3, 4, 7, and 8). B, PKC co-clusters (arrows) with patched syndecan-4 (Syn-4) in spreading cells. Bar = 5 µm. C, immunoblotting with anti-PKC indicates 4V binds PKC and PKM. Material is not released by low salt (lane 1) or EDTA (lane 2), but is by 1 M NaCl (lane 3). No material bound in the absence of calcium and PMA (lane 4). D, solid phase binding assays show minimal binding of recombinant PKCalpha to milk-coated (M) and maximal binding to noncoated (P) wells. Binding increased (p < 0.001, n = 3, bar ± S.E.) to 4V-coated wells when PKCalpha was added in the presence of PL and calcium (compare lanes 4, 5, and 6).
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Fusion Protein Production

cDNA for full-length or truncated core proteins of syndecan-4 in glutathione S-transferase (GST) expression vector pGEX-5X-1 (Pharmacia) were expressed as fusion proteins in Escherichia coli, solubilized with 1% N-laurylsarcosine in 10 mM Tris, pH 7.5, 150 mM NaCl, 2 mM EDTA, 1 mM dithiothreitol, 0.5 mM phenylmethylsulfonyl fluoride, 1 mM benzamidine HCl, purified on glutathione-agarose (Sigma), washed to exchange bound detergent to 0.1% Triton X-100, eluted with 50 mM Tris-HCl, pH 8.0, 5 mM reduced glutathione (Janssen Chimica, New Brunswick, NJ), subjected to 6% SDS-PAGE, and labeled by Coomassie Blue or immunoblotted with anti-GST (Molecular Probes, Eugene, OR).

Phosphorylation Assays

Similar results were seen with PKCalpha beta gamma (1.2 ng) from rat brain (Upstate Biotechnology, Inc.) or 1 ng of recombinant PKCalpha (Molecular Probes and Upstate Biotechnology, Inc.). These were incubated (10 min, room temperature) in 20 µl of 50 mM HEPES, pH 7.3, containing 3 mM magnesium acetate, 750 µM CaCl2, 200 µM ATP (0.5 µCi of [gamma -32P]ATP), 3 µg of histone III-S (Sigma) with preformed phospholipid (PL: 4 µg of PS and 0.4 µg of DL, briefly sonicated) micelles as indicated. Dose-response assays (not shown) determined that these conditions gave maximum activation of PKCalpha . Fusion proteins (1 µg) or synthetic peptides were added to preformed micelles, except for 4L, which was only effective when added during their formation. In phosphorylation assays with protein kinase M (PKM: 1.3 ng; PKC catalytic subunit from rat brain, Calbiochem), calcium was omitted, but PL was present to control for any contamination with whole PKC, which would be activated by PL. The commercial PKM preparation did not contain PKC, since no increased activity was seen on addition of PL (Fig. 3B, compare lanes 1 and 2). Other PKM assays also, therefore, contained PL for comparative purposes. Reactions were stopped by addition of trichloroacetic acid (to 20%) and 0.5% SDS (fusion protein assays) and the precipitates analyzed or by direct analysis following addition of 5 × SDS-PAGE mixture (peptide assays). Products were subjected to 20% SDS-PAGE and autoradiography or PhosphorImager analysis (n = 2-10, values shown are mean ± S.E., means compared by Student's t test). The presence of NH2-terminal cysteine to allow conjugation to affinity matrices had no effect except to enhance direct activation. All assays terminated when activity was maximal and linear.


Fig. 3. Kinase regulation by syndecan-4 V region. A, autoradiographs and quantification show that PKM-mediated phosphorylation of histones was greater in the presence of 1 µg of oligomeric fusion protein 4W than with 4I or 4R (lanes 1-3, n = 2). B, phosphorylation of histones by PKM was enhanced by addition of 50 µg/ml peptides 4L or 4V (lanes 3 and 4, n = 5, p < 0.001) but not the scrambled 4V peptide or 2V (lanes 5 and 6). C, normally maximal PL-mediated phosphorylation of histone by recombinant PKCalpha (lane 2) is potentiated by 50 µg/ml 4V (lane 3, n = 5, p < 0.001). In the absence of PL, PKCalpha is inactive (lane 1), but was directly activated by 250 µg/ml 4V (lane 6, n = 10, p < 0.001). Similar concentrations of scrambled 4V or 2V were ineffective (lanes 4, 5, 7, and 8).
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RESULTS

Syndecan-4 Core Protein Binds PKC and PKM in Vitro and in Vivo

The cytoplasmic domains of the syndecan family members are highly homologous except for one central region (16-18). Since syndecan-4 is the only syndecan present in focal adhesions of fibroblasts, we tested in several ways whether this unique region (here termed 4V) mediated interactions with PKC. First, PKC was co-immunoprecipitated from REF cell lysate with syndecan-4 proteoglycan in the presence, but not the absence, of phorbol ester (PMA) to activate PKC (23). This was prevented by inclusion of the cytoplasmic domain peptide 4V as a competitor, but not by a peptide of scrambled sequence (Fig. 1A). Second, when syndecan-4 was patched on the surface of spreading cells, PKC was co-patched (Fig. 1B). This co-patching of initially diffuse syndecan-4 and PKC was accelerated by PMA (not shown). Third, when cell lysates were passed over an affinity matrix of syndecan 4V peptide (but not the equivalent 2V peptide), two components corresponding in molecular mass to PKCalpha and PKM (the catalytic domain of PKC) were specifically eluted and detected by immunoblotting with antibodies against PKC catalytic domain (Fig. 1C). Again, binding of PKC or PKM was not detected in the absence of calcium and PMA to activate PKC (23) (compare lanes 3 and 4). PMA was the required agent, however, since elution with EDTA (lane 2) had no effect. Fourth, interaction was also seen in solid phase assays (Fig. 1D) where substrates of 4V specifically and directly bound PKCalpha . This assay requires coating substrates with the peptide of interest, adding PKCalpha under differing binding conditions, washing, and then activating bound PKCalpha by addition of PL. Minimal binding occurred to wells coated only with milk (M: negative control; nonspecific binding). Maximal binding occurred to wells blocked with milk after addition of PKCalpha (P: positive control), but phosphorylation was even more evident in wells coated with 4V peptide, since this reflects not just binding, but also potentiation of activity. Binding of PKCalpha was again optimal where the enzyme had been activated with calcium and PL during binding to 4V-coated substrates (compare last three lanes in Fig. 1D).

Syndecan-4 Regulates PKC Activity through Interactions of Its V Region with the Catalytic Domain

Since syndecan-4 appeared to bind PKC, both in vivo and in vitro, we monitored whether this could affect enzyme activity. Fig. 2A shows a diagram of three GST-fusion proteins containing either full-length syndecan-4 core protein (4W) or with truncated cytoplasmic domains (4I, 4R). Fig. 2B shows these purified fusion proteins by Coomassie Blue staining (lanes 1-3) and immunoblotting with antibodies against GST (lanes 4-6). All the fusion proteins migrated with Mr much greater than the predicted molecular mass (44-47 kDa), indicative of the formation of SDS-resistant oligomers as seen in all syndecans and confirming that the cytoplasmic domain is not essential for self-association (24). These fusion proteins were used in in vitro assays to test for regulation of activity of a mixture of PKCalpha beta gamma . When phosphorylation by PKC of histone III-S was already maximal due to the presence of PL and calcium, full-length syndecan-4 fusion protein potentiated the activity of a mixture of PKCalpha beta gamma (lane 1, Fig. 2C) a further 3-fold. This was not seen in the presence of 4I or 4R (Fig. 2C). Similar potentiation was seen with recombinant PKCalpha (not shown). None of the fusion proteins served as a PKC substrate.


Fig. 2. Recombinant syndecan-4 regulates PKC activity. A, GST-fusion proteins (4W, full-length or 4I, 4R, cytoplasmically truncated) and a comparison of the cytoplasmic domains of syndecan-4 and -2. The variable region (V) is noted, together with the peptides synthesized (4L, 4V, 2V). B, Coomassie Blue (lanes 1-3) and anti-GST immunoblots (lanes 4-6) of fusion proteins. C, representative autoradiographs (insets) and quantification show enhanced PL-mediated phosphorylation of histones by a mixture of PKCalpha beta gamma with 1 µg of oligomeric 4W (p < 0.001, n = 5, bar ± S.E.), over that with 1 µg of truncated core proteins 4I and 4R.
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Syndecan-4 appeared to act through the catalytic domain of PKCalpha , since 4W fusion protein, but not truncated fusion protein, could also potentiate the activity of PKM (Fig. 3A), which lacks the regulatory PL- and calcium-binding domains, and is, therefore, constitutively active. It also bound both PKC and PKM (Fig. 1C). From binding studies and the different effects of the truncated fusion proteins, it appeared that the active region in syndecan-4 cytoplasmic domain was the V region. Similar assays were performed, therefore, with synthetic peptides corresponding to the whole cytoplasmic domain (4L) and V (4V, 2V) regions of syndecan-4 or its closest homolog, syndecan-2 (Fig. 3B). PKM is constitutively active in the absence of PL, since it lacks the regulatory domain. An absence of intact PKC in the PKM preparation was confirmed by a comparison of histone phosphorylation in the absence or presence of PL (compare lanes 1 and 2). Peptide 4L (lane 3) and 4V (lane 4) potentiated phosphorylation of histones by PKM with an average increase of 3.5-4-fold. Neither a peptide of scrambled 4V sequence (Scr; lane 5) nor the corresponding sequence from syndecan-2 (2V; lane 6) showed any enhancement of histone phosphorylation. The equivalent sequence from syndecan-1 was also ineffective (not shown). Whole PKCalpha was inactive in the absence of PL (Fig. 3C, lane 1), but activated by PL (lane 2). This PL-mediated PKCalpha activity was potentiated by the addition of peptide 4V (Fig. 3C, lane 3). Quantification indicated an average increase to 302 ± 41% in the presence of 4V over normally maximal PL-mediated activity (Fig. 3C, lane 2), and 40-fold increase over base-line activity (i.e. in the absence of PL; Fig. 3C, lane 1). This was dose-dependent, and clearly detectable at 0.45 µM peptide (not shown). As a further indication of the independent regulatory properties of syndecan-4 V region, histone phosphorylation by PKCalpha was detected, even in the absence of PL (Fig. 3C, lane 6) when 4V was added. Direct activation was also dose-dependent with maximal activation at 0.1 mM (not shown), but never reached the levels seen in the presence of PL; the average increase was 3-fold over base line. No activity was again seen with the scrambled peptide or 2V (Fig. 3C, lanes 7 and 8). Similar potentiation of PL-mediated activation, and direct activation of PKCalpha beta gamma , was seen in assays using a physiological substrate, an epidermal growth factor receptor peptide (25) (not shown). Therefore, the effects of 4V on potentiation of PL-mediated PKC activation were not merely due to facilitation of histone interaction with PL vesicles. Furthermore, direct activation of PKC by 4V peptide can occur in the absence of PL (Fig. 3C, compare lanes 1 and 6).

The 4V region is highly cationic, and protamine has been shown previously to activate PKC in a PL-independent manner (26). However, further experiments with 4V peptides of altered or scrambled sequence confirmed the specificity for PKCalpha regulation by syndecan-4 (Fig. 4, A and B). Even conservative substitutions markedly diminished the ability to potentiate PL-mediated activation or to directly activate PKCalpha in the absence of PL.


Fig. 4. Peptide sequence determines activity. Potentiation of PL-mediated phosphorylation (A) or direct activation (B) of PKCalpha phosphorylation of histones is not enhanced, or is at background levels, if peptides of altered sequence are substituted for 4V. Peptide sequence numbering corresponds to lane numbers on the autoradiographs. Peptides were added at 50 µg/ml (A) or 250 µg/ml (B).
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DISCUSSION

Protein kinase C binding to syndecan-4 was seen using intact core protein and cytoplasmic peptides in vitro and intact glycosylated syndecan-4 in vivo. A specific region of the cytoplasmic domain of syndecan-4 was mapped as a site that interacts with and regulates the activity of PKCalpha and PKM. This region of syndecan-4 is the first example of a protein sequence from a cell surface receptor that can directly regulate PKC localization and activity. Syndecan-4 probably interacts through the catalytic domain of PKC, since it elevated the activity of this isolated domain (PKM), and PKCalpha activity was additive in the presence of both 4V and PL. Binding of PKCalpha to the V region of syndecan-4 in vitro was only clearly detectable after enzyme activation by PMA, whereas co-patching occurred in vivo without additional treatment. Treatment of cells with PMA did, however, accelerate the patching process, indicating that binding of mediators to the regulatory domain may enhance the availability of the catalytic domain of PKC for binding to syndecan-4. PKC is active early during cell spreading (4), and its activation results in syndecan-4 translocation to stabilizing sites of cell-matrix adhesion (20). Therefore, it is possible that, during cell adhesion, conventional activation of PKCalpha facilitates its interaction with ligated and clustered syndecan-4 and the colocalization of this complex to focal adhesions. Syndecan-4 may then, at these sites, compartmentalize PKCalpha and regulate its phosphorylation of other cytoplasmic components.

Consistent with a role for syndecan-4 core protein regulating PKC activity during focal adhesion and stress fiber formation (8-11, 19), we have found2 that the ability of Chinese hamster ovary cells to form these structures is increased after stable transfection with cDNA for syndecan-4 core protein. In contrast, cells transfected with a cDNA encoding a truncated syndecan-4 core protein that terminates in the V region have greatly reduced focal adhesions and stress fibers. Further studies to investigate which cytoskeletal or other cytoplasmic proteins are phosphorylated at adhesions are underway.


FOOTNOTES

*   This work was supported in part by National Institutes of Health Grant GM50194 and the Cell Adhesion and Matrix Research Center at University of Alabama at Birmingham.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Dagger    To whom correspondence should be addressed: Dept. of Cell Biology, Volker Hall 203A, University of Alabama at Birmingham, Birmingham, AL 35294-0019. Tel.: 205-934-2626; Fax: 205-975-9956; E-mail: awoods{at}cellbio.bhs.uab.edu.
1   The abbreviations used are: PKC, protein kinase C; PKM, protein kinase M; DL, diolein; GST, glutathione S-transferase; PL, phospholipid; PMA, phorbol 12-myristate 13-acetate; PS, phosphatidylserine; REF, rat embryo fibroblasts; PAGE, polyacrylamide gel electrophoresis.
2   A. Woods, R. L. Longley, A. Fleetwood, G. Cowling, J. T. Gallagher, and J. R. Couchman, manuscript in preparation.

ACKNOWLEDGEMENTS

We thank the Comprehensive Cancer Center (University of Alabama at Birmingham) for peptide synthesis and analysis, Drs. G. J. Cowling and J. T. Gallagher and A. Fleetwood (University of Manchester, Manchester, United Kingdom) for the syndecan-4 cDNA constructs.


REFERENCES

  1. Miyamoto, S., Teramoto, H., Coso, O. A., Gutkind, J. S., Burbelo, P. D., Akiyama, S. K., and Yamada, K. M. (1995) J. Cell Biol. 131, 791-805 [Abstract]
  2. Clark, E. A., and Brugge, J. S. (1995) Science 268, 233-239 [Medline] [Order article via Infotrieve]
  3. Burridge, K., and Chrzanowska-Wodnicka, M. (1996) Annu. Rev. Cell. Dev. Biol. 12, 463-518 [CrossRef][Medline] [Order article via Infotrieve]
  4. Vuori, K., and Ruoslahti, E. (1993) J. Biol. Chem. 268, 21459-21462 [Abstract/Free Full Text]
  5. Burridge, K., Turner, C. E., and Romer, L. H. (1992) J. Cell Biol. 119, 898-903
  6. Beyth, R. C., and Culp, L. A. (1984) Exp. Cell Res. 155, 537-548 [Medline] [Order article via Infotrieve]
  7. Streeter, H. B., and Rees, D. A. (1987) J. Cell Biol. 105, 507-515 [Abstract]
  8. Woods, A., and Couchman, J. R. (1992) J. Cell Sci. 101, 277-290 [Abstract]
  9. Woods, A., and Couchman, J. R. (1992) Adv. Exp. Med. Biol. 313, 87-96 [Medline] [Order article via Infotrieve]
  10. Woods, A., Couchman, J. R., Johansson, S, and Höök, M. (1986) EMBO J. 5, 665-670 [Abstract]
  11. Woods, A., McCarthy, J. B., Furcht, L. T., and Couchman, J. R. (1993) Mol. Biol. Cell 4, 605-613 [Abstract]
  12. Huhtala, P., Humphries, M. J., McCarthy, J. B., Tremble, P. M., Werb, Z., and Damsky, C. H. (1995) J. Cell Biol. 129, 867-879 [Abstract]
  13. LeBaron, R. G., Esko, J. D., Woods, A., Johansson, S., and Höök, M. (1988) J. Cell Biol. 106, 945-952 [Abstract]
  14. Couchman, J. R., Austria, R., Woods, A., and Hughes, R. C. (1988) J. Cell Physiol. 136, 226-236 [Medline] [Order article via Infotrieve]
  15. Nobes, C. D., and Hall, A. (1995) Cell 81, 53-62 [Medline] [Order article via Infotrieve]
  16. Bernfield, M., Kokenyesi, R., Kato, M., Hinkes, M. T., Spring, J., Gallo, R. L., and Lose, E. J. (1992) Annu. Rev. Cell Biol. 8, 365-393 [CrossRef]
  17. Couchman, J. R., and Woods, A. (1993) in Cell Surface and Extracellular Glycoconjugates (Roberts, D. D., and Mecham, R. P., eds), pp. 33-82, Academic Press, New York
  18. Gallagher, J. T. (1996) in Extracellular Matrix (Comper, W. D., ed), Vol. 2, pp. 230-245, Harwood Academic Press, Amsterdam
  19. Woods, A., and Couchman, J. R. (1994) Mol. Biol. Cell 5, 183-192 [Abstract]
  20. Baciu, P. C., and Goetinck, P. F. (1995) Mol. Biol. Cell 6, 1503-1513 [Abstract]
  21. Hyatt, S. L., Klauck, T., and Jaken, S. (1990) Mol. Carcinogen. 3, 45-53 [Medline] [Order article via Infotrieve]
  22. Burn, P., Kupfer, A., and Singer, S. J. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 497-501 [Abstract]
  23. Gschwendt, M., Kittstein, W., and Marks, F. (1991) Trends Biochem. Sci. 16, 167-169 [CrossRef][Medline] [Order article via Infotrieve]
  24. Asundi, V. K., and Carey, D. J. (1995) J. Biol. Chem. 270, 26404-26410 [Abstract/Free Full Text]
  25. Lund, K. A., Lazar, C. S., Chen, W. S., Walsh, B. J., Welsh, J. B., Herbst, J. J., Walton, G. M., Rosenfeld, M. G., Gill, G. N., and Wiley, H. S. (1990) J. Biol. Chem. 265, 20517-20523 [Abstract/Free Full Text]
  26. Leventhal, P. S., and Bertics, P. J. (1993) J. Biol. Chem. 268, 13906-13913 [Abstract/Free Full Text]

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