A lipid-anchored heparan sulfate proteoglycan is present in the surface of differentiated skeletal muscle cells. Isolation and biochemical characterization.
We have investigated the presence of hydrophobic membrane-associated heparan sulfate proteoglycans (HSPG) on the cell surface of differentiated skeletal muscle cells. A HSPG releasable by incubation with a phosphatidylinositol-specific phospholipase c (PtdIns-PLC) was obtained. HSPG were also isolated from Triton X-100 extracts of the cells. The hydrodynamic characteristics of the PtdIns-PLC-releasable and detergent-extracted HSPG were indistinguishable. SDS/PAGE analysis of the PtdIns-PLC-releasable HSPG indicated a molecular mass of 250 kDa. Analysis of proteins immunoprecipitated by specific antibodies against a HSPG isolated from Schwann cells demonstrated that the antisera precipitated an intact HSPG that was present in the pool of proteins released by PtdIns-PLC and by Triton X-100 from [35S]sulfate labeled cells. Nitrous acid degradation of the immunoprecipitated proteins released by PtdIns-PLC from [35S]methionine labeled cells produced a single 67-kDa core protein. Analysis of hydrophobicity of the purified HSPG revealed that only the HSPG obtained from the detergent extract were able to be incorporated into the liposomes whereas the PtdIns-PLC-released HSPG was not. Immunocytolocalization analysis of the differentiated cells indicated that the PtdIns-PLC-releasable HSPG was located on the cell surface. Prior incubation of the cells with PtdIns-PLC significantly reduced the surface staining. Analysis of skeletal-muscle sections of adult rat skeletal muscles indicated that this HSPG localized exclusively at the endomysium. This localization suggest that these HSPG may be acting as a cell receptor for extracellular-matrix (ECM) components.
Department of Cell and Molecular Biology, Faculty of Biological Sciences, Catholic University of Chile, Santiago.
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A
Campos
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Núñez
C S
CS
Koenig
D J
DJ
Carey
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Brandan
D000818:Animals; D002454:Cell Differentiation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D009132:Muscles / Q000737:chemistry* / Q000166:cytology; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011233:Precipitin Tests; D011509:Proteoglycans / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D012583:Schwann Cells / Q000378:metabolism
D017261:Glycosylphosphatidylinositols; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases; D011509:Proteoglycans