Neurite Outgrowth in Brain Neurons Induced by Heparin-binding Growth-associated Molecule (HB-GAM) Depends on the Specific Interaction of HB-GAM with Heparan Sulfate at the Cell Surface (*)
- Tarja Kinnunen(1),
- Erkki Raulo(1),
- Riitta Nolo(1),
- Marco Maccarana(2),
- Ulf Lindahl(2) and
- Heikki Rauvala(1)(3)
- From the (1)Laboratory of Molecular Neurobiology, Institute of Biotechnology, University of Helsinki, Helsinki 00014, Finland, the
- (2)Department of Medical and Physiological Chemistry, University of Uppsala, The Biomedical Center, S-75123 Uppsala, Sweden, and the
- (3)Department of Biosciences, Division of Biochemistry, University of Helsinki, Helsinki 00014, Finland
Abstract
Heparin-binding growth-associated molecule (HB-GAM) is a cell surface- and extracellular matrix-associated protein that lines developing axons in vivo and promotes neurite outgrowth in vitro. Because N-syndecan (syndecan-3) was found to function as a receptor in HB-GAM-induced neurite outgrowth, we have now studied whether the heparan sulfate side chains of N-syndecan play a role in HB-GAM-neuron interactions. N-Syndecan from postnatal rat brain was found to inhibit HB-GAM-induced but not laminin-induced neurite outgrowth when added to the assay media. The inhibitory activity was abolished by treating N-syndecan with heparitinase, but it was retained in N-syndecan-derived free glycosaminoglycan chains, suggesting that N-syndecan heparan sulfate at the cell surface is involved in HB-GAM-induced neurite outgrowth. Binding to HB-GAM and inhibition of neurite outgrowth was observed with heparin-related polysaccharides only; galactosaminoglycans were inactive. Significant inhibition of neurite outgrowth was induced by heparin and by N-syndecan heparan sulfate but not by heparan sulfates from other sources. A minimum of 10 monosaccharide residues were required for HB-GAM binding, as well as for inhibition of HB-GAM-induced neurite outgrowth. Experiments with selectively desulfated heparins indicated that 2-O-sulfated iduronic acid units, in particular, are of importance to the interaction with HB-GAM, whereas glucosamine N-sulfate and 6-O-sulfate groups were implicated to a lesser extent. Structural analysis of N-syndecan from 6-day-old rat brain indicated that the heparan sulfate chains contain sequences of contiguous, N-sulfated disaccharide units with an unusually high proportion (82%) of 2-O-sulfated iduronic acid residues. We suggest that this property of N-syndecan heparan sulfate is essential for HB-GAM binding and induction of neurite outgrowth.
Footnotes
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↵*This study was supported by grants from the Academy of Finland and from the Sigrid Jusélius Foundation (to H. R.) and by Grant 2309 from the Swedish Medical Research Council (to U. L.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- GAG
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glycosaminoglycan
- HB-GAM
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heparin-binding growth-associated molecule
- HPLC
-
high performance liquid chromatography
- HS
-
heparan sulfate
- aMan
-
anhydromannitol.
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↵2M. Maccarana, Y. Sakura, A. Tawada, K. Yoshida, and U. Lindahl, unpublished observations.
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- Received July 13, 1995.
- Revision received October 31, 1995.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
