Nerve growth factor (NGF) is required for the development of sympathetic neurons and subsets of sensory neurons. Our current knowledge on the molecular mechanisms underlying the biological functions of NGF is in part based on the studies with PC12 rat pheochromocytoma cells, which differentiate into sympathetic neuron-like cells upon NGF treatment. Here we reported that the expression of leukemia inhibitory factor receptor (LIFR), one of the signaling molecules shared by several neuropoietic cytokines of the IL-6 family, was specifically up-regulated in PC12 cells following treatment with NGF. Attenuation of LIFR signaling through stable transfection of antisense or dominant negative LIFR constructs enhanced NGF-induced neurite extension in PC12 cells. On the contrary, overexpression of...[ Read more ]

Nerve growth factor (NGF) is required for the development of sympathetic neurons and subsets of sensory neurons. Our current knowledge on the molecular mechanisms underlying the biological functions of NGF is in part based on the studies with PC12 rat pheochromocytoma cells, which differentiate into sympathetic neuron-like cells upon NGF treatment. Here we reported that the expression of leukemia inhibitory factor receptor (LIFR), one of the signaling molecules shared by several neuropoietic cytokines of the IL-6 family, was specifically up-regulated in PC12 cells following treatment with NGF. Attenuation of LIFR signaling through stable transfection of antisense or dominant negative LIFR constructs enhanced NGF-induced neurite extension in PC12 cells. On the contrary, overexpression of LIFR retarded the growth of neurites. More importantly, while NGF-induced Rac1 activity was enhanced in AS-LIFR and DN-LIFR expressing PC12 cells, it was reduced in LIFR expressing PC12 cells. Following combined treatment with NGF and CNTF, sympathetic neurons exhibited attenuated neurite growth and branching. In sympathetic neurons lacking LIFR, neurite growth and branching was enhanced when compared to wild type controls. Taken together, our findings demonstrate that LIFR expression can be specifically induced by NGF and, besides its known function in cell survival and phenotype development, activated LIFR signaling does exert negative regulatory effect on neurite extension and branching of sympathetic neurons.

Signal transducer and activator of transcription 3 (STAT3) regulates gene transcription in cells in response to cytokines and growth factors. Activation of STAT3 involves phosphorylation of both Tyr705 and Ser727, leading to dimerization, nuclear translocation and DNA binding of STAT3. In the nervous system, STAT3 signaling mediates the differentiation of neuronal and glial cells, as well as survival and phenotypic switching of subtypes of neurons. To further study the mechanism behind the inhibitory effect of LIFR signaling in neurite outgrowth, we have examined the activation of STAT3 during NGF-induced differentiation in PC12 cells. Upon treatment with NGF, phosphorylation of STAT3 on Ser727, but not Tyr705, was detected in PC12 cells. During different stages of neuronal development, phosphorylation of Ser727 by NGF could be mediated through different signaling mechanisms, such as ERK pathway, H7- sensitive signaling pathway, PI3K and/or p35/CDK5 pathway(s). We found that Ser727 phosphorylation of STAT3 induced by NGF could attenuate CNTF-induced Tyr705 phosphorylation in LIFR overexpressing PC12 cells. When Rac1 and STAT3 were overexpressed in COS-7 and PC12 cells, Rac1 was found to associate with STAT3. Both STAT3 and Rac1 could be detected in the growth cones of PC12 cells during the initiation phase of neurite outgrowth. Activation of STAT3 by NGF also enhanced its DNA binding ability and transcription activity. We propose that Ser727 phosphorylation of STAT3 could play an important role in NGF-induced neuronal differentiation.