Molecular dissection of TrkA signal transduction pathways mediating differentiation in human neuroblastoma cells

Activation of the neurotrophin receptor TrkA by its ligand nerve growth fac tor (NGF) initiates a cascade of signaling events leading to neuronal diffe rentiation in vitro and might play an important role in the differentiation of favorable neuroblastomas (NB) in vivo. To study TrkA signal transductio n pathways and their effects on differentiation in NE, we stably expressed wild-type TrkA and five different TrkA mutants in the NGF unresponsive huma n NE cell line SH-SY5Y. Resulting clones were characterized by TrkA mRNA an d protein expression, and by autophosphorylation of the receptor. Introduct ion of wild-type TrkA restored NGF responsiveness of SH-SY5Y cells, as demo nstrated by morphological differentiation, activation of mitogen-activated protein kinases (MAPK) and induction of immediate-early genes, Expression o f TrkA in the absence of NGF resulted in growth inhibition of transfectants compared to parental cells, whereas NGF-treatment increased their prolifer ation rate, Analysis of downstream signal transduction pathways indicated t hat NGF-induced differentiation was dependent on TrkA kinase activity, Our data suggest that several redundant pathways are present further downstream but activation of the RAS/MAPK signaling pathway seems to be of major impo rtance for NGF mediated differentiation of NE cells. Our results also show that the signaling effector SH2-B is a substrate of NGF-mediated Trk signal ing in NE, whereas it is not activated by NGF in rat pheochromocytoma PC12 cells, This might explain the differences we observed in TrkA signaling bet ween neuroblastoma and PC12 cells. Further insight into TrkA signaling may suggest new options for the treatment of NB.