Characterization of functional nerve growth factor-receptors in a CNS glial cell line

Monoclonal antibody 217c recognizes the nerve growth factor-receptor on C6 glioma cells

S. Kumar, J. Huber, L. A. Pena, J. R. Perez-Polo, K. Werrbach-Perez, J. De Vellis

Research output: Contribution to journalArticle

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Abstract

The biological effects of nerve growth factor (NGF) have been shown to be mediated by the high-affinity form of the nerve growth factor receptor (NGF-R) in sympathetic and sensory neurons, and in PC12 cells. We report here that the central nervous system C6 rat glioma cell line likewise expresses functional high-affinity NGF-Rs. The expression of NGF-R mRNA in C6 cells can be up-regulated by cycloheximide and its own ligand, NGF; and it can be rapidly down-regulated by epidermal growth factor (EGF). Furthermore, C6 cells display NGF responsiveness by expressing c-fos mRNA within 30 minutes of treatment with NGF; and after 4-5 days of NGF exposure, C6 cells cease dividing as measured by [3H]-thymidine uptake, change shape, and reveal neurite-like processes. Scatchard analysis of [125I]-labelled NGF bound to solubilized C6 cells confirms the presence of both high- and low-affinity receptor protein. Cross-linking radiolabeled NGF to its receptor in the presence or absence of excess unlabeled NGF, followed by immunoprecipitation with monoclonal antibody (mAb) 192-IgG (a known anti-NGF-R antibody) and SDS-PAGE reveals a 100 kD band corresponding to the NGF/NGF-R complex. An identical band is observed when the immunoprecipitation is carried out with mAb 217c, suggesting that the 217c epitope is related to NGF-R. The 217c antibody was generated against C6 cells and shown to be a cell surface antibody (Peng et al., Science 215:1102-4, 1982); several investigators have used it subsequently as an immunocytochemical marker for Schwann cells. The significance of NGF-Rs in a CNS glial cell line is unclear, but association of NGF with the control of proliferation and/or differentiation of primitive glial cells is suggested.

Original languageEnglish (US)
Pages (from-to)408-417
Number of pages10
JournalJournal of Neuroscience Research
Volume27
Issue number3
DOIs
StatePublished - 1990
Externally publishedYes

Fingerprint

Nerve Growth Factor Receptors
Nerve Growth Factor Receptor
Nerve Growth Factor
Glioma
Neuroglia
Monoclonal Antibodies
Cell Line
Immunoprecipitation
Antibodies
Messenger RNA
PC12 Cells
Schwann Cells

Keywords

  • 192-IgG
  • H-thymidine incorporation
  • c-fos expression
  • C6 glioma
  • High- and low-affinity nerve growth factor-receptor
  • immunoprecipitation
  • mAb 217c

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Characterization of functional nerve growth factor-receptors in a CNS glial cell line : Monoclonal antibody 217c recognizes the nerve growth factor-receptor on C6 glioma cells. / Kumar, S.; Huber, J.; Pena, L. A.; Perez-Polo, J. R.; Werrbach-Perez, K.; De Vellis, J.

In: Journal of Neuroscience Research, Vol. 27, No. 3, 1990, p. 408-417.

Research output: Contribution to journalArticle

Kumar, S. ; Huber, J. ; Pena, L. A. ; Perez-Polo, J. R. ; Werrbach-Perez, K. ; De Vellis, J. / Characterization of functional nerve growth factor-receptors in a CNS glial cell line : Monoclonal antibody 217c recognizes the nerve growth factor-receptor on C6 glioma cells. In: Journal of Neuroscience Research. 1990 ; Vol. 27, No. 3. pp. 408-417.
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AB - The biological effects of nerve growth factor (NGF) have been shown to be mediated by the high-affinity form of the nerve growth factor receptor (NGF-R) in sympathetic and sensory neurons, and in PC12 cells. We report here that the central nervous system C6 rat glioma cell line likewise expresses functional high-affinity NGF-Rs. The expression of NGF-R mRNA in C6 cells can be up-regulated by cycloheximide and its own ligand, NGF; and it can be rapidly down-regulated by epidermal growth factor (EGF). Furthermore, C6 cells display NGF responsiveness by expressing c-fos mRNA within 30 minutes of treatment with NGF; and after 4-5 days of NGF exposure, C6 cells cease dividing as measured by [3H]-thymidine uptake, change shape, and reveal neurite-like processes. Scatchard analysis of [125I]-labelled NGF bound to solubilized C6 cells confirms the presence of both high- and low-affinity receptor protein. Cross-linking radiolabeled NGF to its receptor in the presence or absence of excess unlabeled NGF, followed by immunoprecipitation with monoclonal antibody (mAb) 192-IgG (a known anti-NGF-R antibody) and SDS-PAGE reveals a 100 kD band corresponding to the NGF/NGF-R complex. An identical band is observed when the immunoprecipitation is carried out with mAb 217c, suggesting that the 217c epitope is related to NGF-R. The 217c antibody was generated against C6 cells and shown to be a cell surface antibody (Peng et al., Science 215:1102-4, 1982); several investigators have used it subsequently as an immunocytochemical marker for Schwann cells. The significance of NGF-Rs in a CNS glial cell line is unclear, but association of NGF with the control of proliferation and/or differentiation of primitive glial cells is suggested.

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