Heparan sulfate proteoglycans (HSPGs) synthesized in cultures of rat embryo fibroblasts have been isolated and characterized. Cells grown in the presence of [35S]sulfate secreted a large amount of radiolabeled macromolecules into the culture medium of which only a small proportion was identified as HSPG. However, the majority of radiolabeled proteoglycans isolated from the cell layer were HSPGs. Here, two types of HSPG were detected. One type had an Mr of 5-8 X 10(5) as estimated by gel chromatography on Sepharose CL-4B in the presence of 0.1% sodium dodecyl sulfate and lacked hydrophobic properties in that it showed no affinity for octyl-Sepharose and could not be inserted into liposomes. The other HSPG type had an estimated Mr of 3-5 X 10(5), was retained on octyl-Sepharose, and could be inserted into liposomes. In addition, the cells contained low molecular weight heparan sulfate oligosaccharides. Treatment of living cells with 0.2% Triton X-100, which retained stress fibers and extracellular matrix but solubilized cell membranes, released a proportion of the smaller HSPG together with the heparan sulfate oligosaccharides. The cytoskeleton-matrix residue remaining after detergent extraction of the cell contained the larger species of HSPG in addition to the smaller HSPG. The presence of the smaller hydrophobic HSPG in the detergent-treated cytoskeleton-matrix preparations suggests that it may form part of a transmembrane cytoskeleton-matrix linkage.
Keywords: Animals; Cells, Cultured; Centrifugation, Density Gradient; Cytoskeleton; Embryo, Mammalian; Fibroblasts; Glycosaminoglycans; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Liposomes; Microscopy, Electron, Scanning; Proteochondroitin Sulfates; Proteoglycans; Rats