LYSOSOMAL SULFATE EFFLUX FOLLOWING GLYCOSAMINOGLYCAN DEGRADATION - MEASUREMENTS IN ENZYME-SUPPLEMENTED MAROTEAUX-LAMY SYNDROME FIBROBLASTS AND ISOLATED LYSOSOMES

Studies using lysosomal membrane vesicles have suggested that efflux o f the sulfate that results from lysosomal glycosaminoglycan degradatio n is carrier-mediated. In this study, glycosaminoglycan degradation an d sulfate efflux were examined using cultured skin fibroblasts and lys osomes deficient in the lysosomal enzyme N-acetylgalactosamine-4-sulfa tase. Such fibroblasts store dermatan sulfate lysosomally, which could be labelled biosynthetically with (Na2SO4)-S-35. The addition of reco mbinant N-acetylgalactosamine-4-sulfatase to the media of S-35 labelle d fibroblasts degraded up to 82% of the stored dermatan [S-35] sulfate over a subsequent 96 h chase and released inorganic [S-35] sulfate in to the medium. In the presence of cetamido-4'-isothiocyanatostilbene-2 ,2'-disulfonic acid (SITS), sulfate was reused to a minor extent in ne wly synthesized proteoglycan. Isolated granules from recombinant enzym e supplemented fibroblasts degraded stored dermatan [S-35]sulfate to s ulfate which was rapidly released into the medium at a rate that was r educed by the extra-lysosomal presence of the lysosomal sulfate transp ort inhibitors SITS, Na2SO4 and Na2MoO4. SITS also inhibited dermatan sulfate turnover, although it had no effect on the action of purified recombinant enzyme in vitro. These data imply that sulfate clearance o ccurred concomitantly with dermatan sulfate turnover in the lysosome e ven at high substrate loading, and that lysosome-derived sulfate, whil e available, is reutilized minimally in synthetic pathways.