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Lowe (oculocerebrorenal) syndrome (LS) is an X-linked disorder characterized by congenital cataracts, generalized hypotonia, mental retardation, and renal Fanconi syndrome. The basic defect remains unknown, but the possibility that fibroblasts express reduced sulfation of glycosaminoglycans has been studied in several laboratories. A mechanism involving overproduction of an enzyme (nucleotide pyrophosphatase) active against adenosine 3′-phosphate, 5′-phosphosulfate (PAPS) has been postulated. Decreased synthesis of normally sulfated glycosaminoglycans was also reported. We measured the synthesis of proteoglycans and glycosaminoglycans by incorporation of [3H]glucosamine and Na2(35)SO4 into cultured fibroblasts from four LS patients and related it directly to the synthesis in six normal fibroblast cultures. We found that the rate of synthesis varied greatly among the normal cultures (cv, 30%), but not significantly between LS and the normal. The LS fibroblasts' ability to sulfate glycosaminoglycans was assayed as the amount of 3H-glycosaminoglycan eluting at low ionic strength on anion exchange chromatography, the amount of non-sulfated disaccharide present in chondroitinase digests of labeled proteoglycans, and the ratio of 35S to 3H incorporation into proteoglycans. Each parameter suggested that the LS cells were synthesizing normally sulfated glycosaminoglycans (e.g. % delta Di-0S, 21 +/- 6 in normal; 27 +/- 6 in LS). The cells' ability to sulfate glycosaminoglycans was tested under conditions of markedly stimulated glycosaminoglycan synthesis, by treating the cultures with a beta-D-xyloside. LS and normal cells responded to the treatment by elevating the rate of synthesis of normally sulfated glycosaminoglycans (3.5-6-fold in normal, 3-7-fold in LS). Nucleotide pyrophosphatase activities were found to be elevated in each of our four LS cell strains as in the previous studies, excluding genetic heterogeneity as an explanation for our findings. We conclude that LS fibroblasts do not express defects in sulfation of glycosaminoglycans or in synthesis of proteoglycans.

Percoll-purified rat thyroid FRTL-5 cell lysosomes were photoaffinity-labeled with [125I]diiodotyrosine to identify proteins which bind diiodotyrosine, a ligand for lysosomal transport system h. SDS-PAGE and autoradiography of these membranes showed specific labeling of a 70-kDa protein and weak labeling of three smaller proteins. [125I]Diiodotyrosine photolabeling of the 70-kDa protein was specifically competed against by ligands of lysosomal transport system h ligands. The 70-kDa protein was photolabeled more strongly in lysosomal membranes isolated from thyrotropin-stimulated cells when compared with those grown in the absence of thyrotropin, consistent with previous demonstrations that thyrotropin stimulates system h transport. The 70-kDa protein may represent some portion of the system h carrier protein.Copyright 1995, 1999 Academic Press, Inc.