Release of the NILE and Other Glycoproteins from Cultured PC 12 Rat Pheochromocytoma Cells and Sympathetic Neurons

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Abstract
Studies were carried out on the glycoproteins (GPs) released by cultured rat sympathetic neurons and by cultured PC12 rat pheochromocytoma cells with and without nerve growth factor (NGF) treatment. Cultures were prelabeled with [3H]fucose and then incubated for 4‐8 h in fresh unlabeled medium. The material released into the medium was analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and fluorography. The patterns of labeled material released by all three types of cultures were similar. One of the major components released was of apparent Mr≲ 230,000. Another major component of apparent Mr= 55,000 as well as minor components of apparent Mr≲ 180,000, 140,000, 118,000, and 105,000 were also detected. An additional peptide of apparent Mr≲ 210,000 was released only by the sympathetic neurons. The soluble released Mr≲ 230,000 component appeared to be derived from a previously characterized neuronal integral membrane GP referred to as the NILE (NGF‐inducible large external) GP. Evidence for this included recognition of the released component by a monospecific antiserum prepared against membrane‐derived NILE GP. At least several of the other released GPs appeared to be derived from membrane‐bound components with which they share immuno‐crossreactivity. Since the soluble NILE and other released GPs had somewhat faster mobilities on SDS‐polyacrylamide gels than their apparent membrane‐bound correspondents, release could either be due to, or accompanied by, minor changes in molecular structure.