Low Buoyant Density Proteoglycans from Saline and Dissociative Extracts of Embryonic Chicken Retinas

Abstract

Retinas were labeled in culture with [3H]glucosamine or [3H]leucine and [35S]sulfate and extracted sequentially with physiologically balanced saline and 4 M guanidine HCl. They were dialyzed into associative conditions (0.5 M NaCl) and chromatographed on agarose columns. Under these conditions, some of the proteoglycans were associated in massive complexes that showed low buoyant densities when centrifuged in CsCl density gradients under dissociative conditions (4 M guanidine HCl). Much of the label in these complexes was in molecules other than proteoglycans. Most of the proteoglycans, however, were included on the agarose columns, where they appeared to be constitutionally of low buoyant density. They resisted attempts to separate potential low buoyant density contaminants from the major proteoglycans by direct CsCl density gradient centrifugation or by the fractionation of saline or 8 M urea extracts on diethylaminoethyl-Sephacel. The diethylaminoethyl-Sephacel fractions were either subjected to CsCl density gradient centrifugation or were chromatographed on Sephacryl S-300, in both cases before and after alkaline cleavage, to confirm the presence of typical O-linked glycosaminoglycans. The medium and balanced salt extracts were enriched in chondroitin sulfate and other sulfated macromolecules, possibly highly sulfated oligosaccharides, that resisted digestion by chondroitinase ABC but were electrophoretically less mobile than heparan sulfate. Guanidine HCl or urea extracts of the residues were mixtures of high and low density proteoglycans that were enriched in heparan sulfate.

Transient Expression of PG‐M/Versican, a Large Chondroitin Sulfate Proteoglycan in Developing Chicken Retina

“…Several chondroitin sulfate proteoglycans have been identified and characterized in the nervous system (for review, see Small et al, 1996) and in the retina (Morris and Ting, 1981;Aquino et al, 1984;Morris, 1984;Morris et al, 1984Morris et al, , 1987Needham et a!., 1988;Brittis et al, 1992;Ring et a!., 1995 to extracellular matrix molecules, such as fibronectin, and that the activity can be attributed to chondroitin sulfate chains (Yamagata et al, 1989). Indeed, PG-MI versican-like chondroitin sulfate proteoglycan purified from sciatic nerve inhibited adhesion of several nerve cells and affected neurite outgrowth (Braunewell et al, 1995).…”

Axonal transport of proteoglycans to the goldfish optic tectum

“…This has included the isolation of molecules containing chondroitin sulfate from rat brain (1), retina (2,3), and cultured glial cells (4). Heparan sulfate proteoglycans have been isolated from the same sources (2,(5)(6)(7)(8) as well as from calf brain micro-somes (9), cultured PC 12 cells (10), and neuroblastoma cells (11).…”