Fast and slow nerve growth factor binding sites in human neuroblastoma and rat pheochromocytoma cell lines: relationship of sites to each other and to neurite formation

• Published in: The Journal of neuroscience 1985-07, Vol.5 (7), p.1717-1728
• Main Authors: Sonnenfeld, KH, Ishii, DN
• Format: Article
• Language: English
• Subjects: Animals; Binding Sites; Biological and medical sciences; Cell Line; Male; Medical sciences; Mice; Mice, Inbred Strains; Nerve Growth Factors - metabolism; Nerve Growth Factors - physiology; Neuroblastoma - metabolism; Neurology; Neurons - growth & development; Neurons - metabolism; Pheochromocytoma - metabolism; Sympathetic Nervous System - metabolism; Sympathetic Nervous System - physiology; Tumors of the nervous system. Phacomatoses
• ISSN: 0270-6474; 1529-2401
• DOI: 10.1523/jneurosci.05-07-01717.1985

We studied (a) the distribution and properties of fast and slow 125I-nerve growth factor (125I-NGF) binding sites in cultured human neuroblastoma (NB) cell lines that were categorized as responsive (N+) or unresponsive (N-) to NGF by neurite outgrowth, (b) whether fast or slow sites mediate actions of NGF, and (c) whether NGF-mediated conversion of fast to slow sites occurs in human NB and pheochromocytoma PC 12 cells. In human NB SH-SY5Y cells, the slow sites were trypsin resistant and binding was of high affinity. Loss of binding to the slow sites had a half-time of 25 to 30 min at 37 degrees C and was very slow at 4 degrees C. In contrast, the fast sites were trypsin sensitive and binding was of lower affinity; its dissociation half-time was less than 1 min at 4 degrees C and 37 degrees C. The association rate constants of both sites were about 0.8 to 1.2 X 10(7) M-1 sec-1. Some human NB cells had both fast and slow sites. The N+ human NB lines SH-SY5Y and LA-N-5 had only slow sites. Despite the virtual elimination of fast sites by trypsin in NB MC-IXC cells, remaining slow sites could still efficiently bind 125I-NGF. These observations showed that fast sites are not required for slow site binding, neurite outgrowth, or other demonstrated actions of NGF in some NB cells. In PC 12 cells, 125I-NGF initially bound to fast sites was not directly transferred to slow sites as required for NGF-mediated conversion. The association rate constants of fast and slow sites in PC12 cells were both about 2 X 10(7) M-1 sec-1. The association kinetics were consistent with simple bimolecular reactions in both NB and PC12 cells. The combined evidence in NB and PC12 cells did not support the hypothesis of NGF-mediated conversion of fast to slow sites.