Activation of nuclear factor κB and bcl-x survival gene expression by nerve growth factor requires tyrosine phosphorylation of IκBα

NGF has been shown to support neuron survival by activating the transcription factor nuclear factor-kappaB (NF kappaB). We investigated the effect of NGF on the expression of Bcl-xL, an anti-apoptotic Bcl-2 family protein. Treatment of rat pheochromocytoma PC12 cells, human neuroblastoma SH-SY5Y cells, or primary rat hippocampal neurons with NGF (0.1-10 ng/ml) increased the expression of bcl-xL mRNA and protein. Reporter gene analysis revealed a significant increase in NF kappaB activity after treatment with NGF that was associated with increased nuclear translocation of the active NF kappaB p65 subunit. NGF-induced NF kappaB activity and Bcl-xL expression were inhibited in cells overexpressing the NF kappaB inhibitor, I kappaB alpha. Unlike tumor necrosis factor-alpha (TNF-alpha), however, NGF-induced NF kappaB activation occurred without significant degradation of I kappa Bs determined by Western blot analysis and time-lapse imaging of neurons expressing green fluorescent protein-tagged I kappaB alpha. Moreover, in contrast to TNF-alpha, NGF failed to phosphorylate I kappaB alpha at serine residue 32, but instead caused significant tyrosine phosphorylation. Overexpression of a Y42F mutant of I kappaB alpha potently suppressed NFG-, but not TNF-alpha -induced NF kappaB activation. Conversely, overexpression of a dominant negative mutant of TNF receptor-associated factor-6 blocked TNF-alpha-, but not NGF-induced NF kappaB activation. We conclude that NGF and TNF-alpha induce different signaling pathways in neurons to activate NF kappaB and bcl-x gene expression.