The Journal of Cell Biology, Vol 111, 2053-2062, Copyright © 1990 by The Rockefeller University Press

ARTICLES

Identification of a lipid-anchored heparan sulfate proteoglycan in Schwann cells

DJ Carey and RC Stahl
Weis Center for Research, Geisinger Clinic, Danville, Pennsylvania 17822.

Schwann cells synthesize both hydrophobic and peripheral cell surface heparan sulfate proteoglycans (HSPGs). Previous analysis of the kinetics of radiolabeling suggested the peripheral HSPGs are derived from the membrane-anchored forms (Carey, D., and D. Evans. 1989. J. Cell Biol. 108:1891-1897). Peripheral cell surface HSPGs were purified from phytic acid extracts of cultured neonatal rat sciatic nerve Schwann cells by anion exchange, gel filtration, and laminin-affinity chromatography. Approximately 250 micrograms of HSPG protein was obtained from 2 X 10(9) cells with an estimated recovery of 23% and an overall purification of approximately 2000-fold. SDS-PAGE analysis indicated the absence of non-HSPG proteins in the purified material. Analysis of heparinase digestion products revealed the presence of at least six core protein species ranging in molecular weight from 57,000 to 185,000. The purified HSPGs were used to produce polyclonal antisera in rabbits. The antisera immunoprecipitated a subpopulation of 35SO4- labeled HSPGs that were released from Schwann cells by incubation in medium containing phosphatidylinositol-specific phospholipase C (PI- PLC); smaller amounts of immunoprecipated HSPGs were also present in phytic acid extracts. In the presence of excess unlabeled PI-PLC- released proteins, immunoprecipitation of phytic acid-solubilized HSPGs was inhibited. SDS-PAGE analysis of proteins immunoprecipitated from extracts of [35S]methionine labeled Schwann cells demonstrated that the antisera precipitated an HSPG species that was present in the pool of proteins released by PI-PLC, with smaller amounts present in phytic acid extracts. Nitrous acid degradation of the immunoprecipitated proteins produced a single 67,000-Mr core protein. When used for indirect immunofluorescence labeling, the antisera stained the external surface of cultured Schwann cells. Preincubation of the cultures in medium containing PI-PLC but not phytic acid significantly reduced the cell surface staining. The antisera stained the outer ring of Schwann cell membrane in sections of adult rat sciatic nerve but did not stain myelin or axonal membranes. This localization suggests the HSPG may play a role in binding the Schwann cell plasma membrane to the adjacent basement membrane surrounding the individual axon-Schwann cell units.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Facebook    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

• Chernousov, M. A., Rothblum, K., Stahl, R. C., Evans, A., Prentiss, L., Carey, D. J. (2006). Glypican-1 and {alpha}4(V) Collagen Are Required for Schwann Cell Myelination. J. Neurosci. 26: 508-517 [Abstract] [Full Text]  
• Li, S., Liquari, P., McKee, K. K., Harrison, D., Patel, R., Lee, S., Yurchenco, P. D. (2005). Laminin-sulfatide binding initiates basement membrane assembly and enables receptor signaling in Schwann cells and fibroblasts. JCB 169: 179-189 [Abstract] [Full Text]  
• Rothblum, K., Stahl, R. C., Carey, D. J. (2004). Constitutive Release of {alpha}4 Type V Collagen N-terminal Domain by Schwann Cells and Binding to Cell Surface and Extracellular Matrix Heparan Sulfate Proteoglycans. J. Biol. Chem. 279: 51282-51288 [Abstract] [Full Text]  
• Erdman, R., Stahl, R. C., Rothblum, K., Chernousov, M. A., Carey, D. J. (2002). Schwann Cell Adhesion to a Novel Heparan Sulfate Binding Site in the N-terminal Domain of alpha 4 Type V Collagen Is Mediated by Syndecan-3. J. Biol. Chem. 277: 7619-7625 [Abstract] [Full Text]  
• Chernousov, M. A., Stahl, R. C., Carey, D. J. (2001). Schwann Cell Type V Collagen Inhibits Axonal Outgrowth and Promotes Schwann Cell Migration via Distinct Adhesive Activities of the Collagen and Noncollagen Domains. J. Neurosci. 21: 6125-6135 [Abstract] [Full Text]  
• Valles, S., Tsoi, C., Huang, W.-Y., Wyllie, D., Carlotti, F., Askari, J. A., Humphries, M. J., Dower, S. K., Qwarnstrom, E. E. (1999). Recruitment of a Heparan Sulfate Subunit to the Interleukin-1 Receptor Complex. REGULATION BY FIBRONECTIN ATTACHMENT. J. Biol. Chem. 274: 20103-20109 [Abstract] [Full Text]  
• Schroering, A., Carey, D. J. (1998). Sensory and Motor Neuron-derived Factor Is a Transmembrane Heregulin That Is Expressed on the Plasma Membrane with the Active Domain Exposed to the Extracellular Environment. J. Biol. Chem. 273: 30643-30650 [Abstract] [Full Text]  
• Rahmatullah, M., Schroering, A., Rothblum, K., Stahl, R. C., Urban, B., Carey, D. J. (1998). Synergistic Regulation of Schwann Cell Proliferation by Heregulin and Forskolin. Mol. Cell. Biol. 18: 6245-6252 [Abstract] [Full Text]  
• Carey, D. J., Bendt, K. M., Stahl, R. C. (1996). The Cytoplasmic Domain of Syndecan-1 Is Required for Cytoskeleton Association but Not Detergent Insolubility. IDENTIFICATION OF ESSENTIAL CYTOPLASMIC DOMAIN RESIDUES. J. Biol. Chem. 271: 15253-15260 [Abstract] [Full Text]  
• Chernousov, M. A., Stahl, R. C., Carey, D. J. (1996). Schwann Cells Secrete a Novel Collagen-like Adhesive Protein That Binds N-Syndecan. J. Biol. Chem. 271: 13844-13853 [Abstract] [Full Text]  
• McCarthy, J., Turley, E.A. (1993). Effects of Extracellular Matrix Components on Cell Locomotion. CROBM 4: 619-637 [Abstract] [Full Text]  
• Chernousov, M. A., Rothblum, K., Tyler, W. A., Stahl, R. C., Carey, D. J. (2000). Schwann Cells Synthesize Type V Collagen That Contains a Novel alpha 4 Chain. MOLECULAR CLONING, BIOCHEMICAL CHARACTERIZATION, AND HIGH AFFINITY HEPARIN BINDING OF alpha 4(V) COLLAGEN. J. Biol. Chem. 275: 28208-28215 [Abstract] [Full Text]  

Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents