Turnover of heparan sulfate depends on 2-O-sulfation of uronic acids.

To study how the pattern of sulfation along a heparan sulfate chain affects its turnover, we examined heparan sulfate catabolism in wild-type Chinese hamster ovary cells and mutant pgsF-17, defective in 2-O-sulfation of uronic acid residues (Bai, X., and Esko, J. D. (1996) J. Biol. Chem. 271, 17711-17717). Heparan sulfate from the mutant contains normal amounts of 6-O-sulfated glucosamine residues and iduronic acid and somewhat higher levels of N-sulfated glucosamine residues but lacks any 2-O-sulfated iduronic or glucuronic acid residues. Pulse-chase experiments showed that both mutant and wild-type cells transport newly synthesized heparan sulfate proteoglycans to the plasma membrane, where they shed into the medium or move into the cell through endocytosis. Internalization of the cell-associated molecules leads to sequential endoglycosidase (heparanase) fragmentation of the chains and eventual lysosomal degradation. In wild-type cells, the chains begin to degrade within 1 h, leading to the accumulation of intermediate (10-20-kDa) and small (4-7-kDa) oligosaccharides. Mutant cells did not generate these intermediates, although internalization and intracellular trafficking of the heparan sulfate chains appeared normal, and the chains degraded with normal kinetics. This difference was not due to defective heparanase activities in the mutant, since cytoplasmic extracts from mutant cells cleaved wild-type heparan sulfate chains in vitro. Instead, the heparan sulfate chains from the mutant were relatively resistant to degradation by cellular heparanases. These findings suggest that 2-O-sulfated iduronic acid residues in heparan sulfate are important for cleavage by endogenous heparanases but not for the overall catabolism of the chains.