Ammonium chloride interferes with a distinct step in the biosynthesis and cell surface expression of human melanoma-type chondroitin sulfate proteoglycan.
Human melanoma cells synthesize a cell-associated chondroitin sulfate-rich proteoglycan, whose core protein is recognized by monoclonal antibody 9.2.27. We report that the core protein is present on the surface of melanoma cells in two forms, either free or modified by the addition of chondroitin sulfate chains, suggesting that the addition of glycosaminoglycan chains may not be a prerequisite for cell surface expression of the proteoglycan core protein. Free core protein found at the cell surface does not seem to represent an overflow of the proteoglycan synthetic pathway, since experiments using a beta-D-xyloside acceptor suggest that core protein is, in fact, limiting proteoglycan synthesis. NH4Cl inhibits the synthesis of melanoma-type proteoglycan, shifting the balance of surface core protein toward the free form. The inhibition of proteoglycan synthesis is apparently not due to a disruption of enzymes and precursors involved in glycosaminoglycan synthesis, since cells treated with NH4Cl retain their ability to initiate and elongate chondroitin 4-sulfate chains on a beta-D-xyloside acceptor. In contrast, the divalent ionophore monensin inhibited core protein maturation and synthesis of glycosaminoglycan chains. The effects of both NH4Cl and monensin were reversible; thus, experiments using the drugs sequentially indicated that monensin temporally precedes NH4Cl in interfering with proteoglycan biosynthesis. Since the NH4Cl and monensin share the property of inhibiting the acidification of intracellular vesicles within cells, the present findings raise the possibility that the accessibility of proteoglycan core protein to the Golgi site of glycosaminoglycan addition is regulated in melanoma cells by acidification of intracellular compartments.
D053674:Aggrecans; D000643:Ammonium Chloride / Q000494:pharmacology*; D011508:Chondroitin Sulfate Proteoglycans / Q000096:biosynthesis*; D016326:Extracellular Matrix Proteins; D005944:Glucosamine / Q000378:metabolism; D006023:Glycoproteins / Q000096:biosynthesis; D006025:Glycosaminoglycans / Q000096:biosynthesis; D006801:Humans; D066298:In Vitro Techniques; D037181:Lectins, C-Type; D008545:Melanoma / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology; D011509:Proteoglycans / Q000096:biosynthesis*; D014316:Tritium
D053674:Aggrecans; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006025:Glycosaminoglycans; D037181:Lectins, C-Type; D011509:Proteoglycans; D000643:Ammonium Chloride; D014316:Tritium; D008985:Monensin; D005944:Glucosamine