INDUCTION OF NERVE GROWTH-FACTOR RESPONSIVENESS IN C6-2B GLIOMA-CELLSBY EXPRESSION OF TRKA PROTOONCOGENE

Cells that lack the high affinity receptor component (trkA) for nerve growth factor (NGF) are unresponsive to NGF. We investigated whether C 6-2B cells, a rat glioma derived cell line, express trkA and, as a con sequence, are responsive to NGF. In these cells, NGF (100 ng/ml) faile d to induce the mRNA encoding for c-fos protooncogene and the low affi nity NGF receptor p75(NGFR), two NGF-responsive genes. In contrast, bo th mRNAs were induced in PC12 cells by NGF. Using a RNase protection a ssay with a cRNA probe for rat trkA, the expected trkA RNA protected f ragment 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 I-125-labeled NGF to PC12 cells identif ied two major bands with an apparent molecular weight of 158 kDa and 1 00 kDa corresponding to trkA and p75(NGFR), respectively. In contrast, only the 100 kDa band could be detected in C6-2B cells by cross-linki ng analysis. In C6-2B cells stably transfected with the rat trkA cDNA, NGF increased c-fos mRNA, induced tyrosine phosphorylation of gp140(t rk), and SNT (suc-associated neurotrophic factor-induced tyrosine-phos phorylated 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 trkA exp ression. Most important, in C6trk(+) cells, NGF was a weaker (2-fold) inducer of [H-3]thymidine incorporation when compared to bFGF (5-fold) , suggesting that expression of trkA fails to confer to NGF a strong m itogenic effect. Our findings indicate that C6-2B glioma cells do not possess high affinity NGF receptor and thus are unresponsive to NGF an d that expression of trkA in neuroectoderm derived cells elicits some of the NGF responses characteristic of neuronal cells. (C) 1994 Wiley- Liss, Inc.