Induction of nerve growth factor responsiveness in C6‐2B glioma cells by expression of trk A proto‐oncogene

Cells that lack the high affinity receptor component ( trk A) for nerve growth factor (NGF) are unresponsive to NGF. We investigated whether C6‐2B cells, a rat glioma derived cell line, express trk A and, as a consequence, are responsive to NGF. In these cells, NGF (100 ng/ml) failed to induce the mRNA encoding for c‐fos protooncogene and the low affinity NGF receptor p75 NGFR , two NGF‐responsive genes. In contrast, both mRNAs were induced in PC12 cells by NGF. Using a RNase protection assay with a cRNA probe for rat trk A RNA protected fragment was detected in PC12 but not in C6‐2B glioma cells, indicating that C6‐2B cells either do not express the gene or express it only in low amounts. Cross‐linking of 125 I‐labeled NGF to PC12 cells identified two major bands with an apparent molecular weight of 158 kDa and 100 kDa corresponding to trk A and p75 NGFR , respectively. In contrast, only the 100 kDa band could be detected in C6‐2B cells by cross‐linking analysis. In C6‐2B cells stably transfected with the rat trk A cDNA, NGF increased c‐fos mRNA, induced tyrosine phosphorylation of gp140 trk , and SNT ( suc ‐associated neurotrophic factor‐induced tyrosinephosphorylated target), and caused morphological changes within 72 h. All of these effects of NGF were blocked by the protein kinase inhibitor K‐252a suggesting that NGF signal transduction was restored by trk A expression. Most important, in C6trk + cells, NGF was a weaker (2‐fold) inducer of [ 3 H]thymidine incorporation when compared to bFGF (5‐fold), suggesting that expression of trk A fails to confer to NGF a strong mitogenic effect. Our findings indicate that C6‐2B glioma cells do not possess high affinity NGF receptor and thus are unresponsive to NGF and that expression of trk A in neuroectoderm derived cells elicits some of the NGF responses characteristic of neuronal cells.