A novel role for nitric oxide in the endogenous degradation of heparan sulfate during recycling of glypican-1 in vascular endothelial cells

We show here that the endothelial cell-line ECV 304 expresses the heparan s ulfate proteoglycan glypican-1. The predominant cellular glycoform carries truncated side-chains and is accompanied by heparan sulfate oligosaccharide s. Treatment with brefeldin A results in accumulation of a glypican proteog lycan with full-size side-chains while the oligosaccharides disappear. Duri ng chase the glypican proteoglycan is converted to partially degraded hepar an sulfate chains and chain-truncated proteoglycan, both of which can be ca ptured by treatment with suramin. The heparan sulfate chains in the intact proteoglycan can be depolymerized by nitrite-dependent cleavage at internal ly located N-unsubstituted glucosamine moieties. Inhibition of NO-synthase or nitrite-deprivation prevents regeneration of intact proteoglycan from tr uncated precursors as well as formation of oligosaccharides. In nitrite-dep rived cells, formation of glypican proteoglycan is restored when NO-donor i s supplied. We propose that, in recycling glypican-1, heparan sulfate chain s are cleaved at or near glucosamines with unsubstituted amino groups. NO-d erived nitrite is then required for the removal of short, nonreducing termi nal saccharides containing these N-unsubstituted glucosamine residues from the core protein stubs, facilitating re-synthesis of heparan sulfate chains .