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The small leucine-rich proteoglycan (SLRP) decorin is efficiently internalized by a variety of cultured cells. A 51-kDa protein has previously been described as a receptor mediating endocytosis of decorin and of the structurally related SLRP biglycan. Recent findings suggest that endocytosis of SLRPs may also be mediated by additional receptors. The class-A scavenger receptor, the endocytic mannose receptor, the epidermal growth factor receptor, and insulin-like growth factor-I receptor have emerged as candidates. We used a combined approach of immunoprecipitation and photoactivated cross-linking to identify endocytosis receptors for decorin in human skin fibroblasts. Decorin was purified by HPLC-DEAE-ion exchange chromatography from the secretions of human skin fibroblasts under nondenaturing conditions. Confocal immunofluorescence microscopy revealed that both biotinylated decorin and decorin conjugated to the heterobifunctional cross-linker sulfosuccinimidyl 2-(p-azidosalicylamido)ethyl-1-3'-dithiopropionate (SASD) were endocytosed with equal efficiency. SASD-conjugated decorin was added to [35S]-methionine-labeled fibroblasts and cross-linked intracellularly to receptor molecules by photoactivation on endocytic uptake. Cross-linked decorin-receptor complexes were purified from the extracts of trypsin-treated fibroblasts by anion exchange chromatography and immunoprecipitation with a decorin-specific antiserum. Analysis by 2D electrophoresis and autoradiography revealed that decorin was specifically cross-linked to a protein of 110 kDa, which exhibited an isoelectric point of 5.5. In a second approach, unlabeled fibroblasts were subjected to decorin endocytosis and photoactivated cross-linking followed by Western blotting of DEAE-purified cell extracts. A shift of biotinylated decorin immunoreactivity from 165 kDa (decorin-receptor complex) to 54 kDa (SASD-conjugated biotinylated decorin) was noted on reductive cleavage of the cross-linker, representing a difference in molecular weight of approximately 110 kDa. The identification of a 110-kDa protein as a novel endocytosis receptor for decorin provides further support for the emerging concept of a redundancy of receptor molecules in the endocytosis of SLRP.

Bisdermican (PG760) is a large, heterodimeric, dermatan sulfate proteoglycan found in selected basement membranes, smooth muscle cell layers, and different extracellular matrices. Age-dependent and developmentally regulated alterations in glycosaminoglycan structure and quantity have been shown to be functionally relevant for a number of physiological and pathological processes. Bisdermican was purified from human skin fibroblast cultures of different age and confluency. Following β-elimination, glycosaminoglycan chains were analyzed by Sephacryl-S-300 chromatography. Glycosaminoglycan chains of Bisdermican from infantile fibroblasts had a molecular weight of 19 kDa, whereas the glycosaminoglycan chain of the large Bisdermican subunit purified from confluent fetal fibroblast secretions was slightly larger (M r = 24 kDa). Bisdermican derived from subconfluent cultures of fetal fibroblasts displayed the largest glycosaminoglycan chains with a molecular weight of 31.5 kDa for the large subunit, and a molecular weight of 22 kDa for the small subunit. Thus, Bisdermican displays a molecular polymorphism that is related to its chronological age and proliferative state. KEYWORDS: glycosaminoglycan, dermatan sulfate, developmental regulation, aging, glycosylation DOMAINS: biochemistry, glycoscience, extracellular matrix, cell biology, cell and tissue culture, aging INTRODUCTIONProteoglycans are a class of glycoproteins of the extracellular matrix and cell surfaces that are characterized by the presence of one or several glycosaminoglycan chains, which are covalently attached to a core protein. Glycosaminoglycans are long, unbranched, anionic polysaccharides containing repetitive disaccharide units. These disaccharide units contain two modified carbohydrates. With the exception of the galactose-containing keratan sulfate, N-acetylgalactosamine or N-acetylglucosamine is linked to a uronic acid such as glucuronic acid or iduronic acid [1,2,3]. Depending on the disaccharide composition and the degree of epimerization and sulfation, dermatan/chondroitin sulfate, heparan D-48149 Münster, Germany; Tel: (+49) 251 56113/Fax: (+49) 251 8356114 **Deceased. ©2004 with author. TheScientificWorldJOURNAL (2004TheScientificWorldJOURNAL ( ) 4, 1017TheScientificWorldJOURNAL ( -1026 sulfate/heparin, keratan sulfate, and hyaluronic acid can be distinguished. Proteoglycans play major roles in a variety of physiological processes and are no longer only considered structural components. For example, proteoglycans have been implicated in the regulation of cell adhesion, migration, differentiation, and signaling in inflammation, development, wound repair, and tumorigenesis [1,2,3,4,5,6]. While some proteoglycan functions are mediated by the core protein, a major part is mediated by the glycosaminoglycans, which can form ligand binding sites defined by the degree of epimerization and sulfation, and the organization into domain structures [2]. Götte et al.: Molecular polymorphism of BisdermicanChanges in the content and composition of glycosaminog...