Abstract
Differentiation of PC12 cells by nerve growth factor (NGF) is characterized by changes in signal transduction pathways leading to growth arrest and neurite extension. The transcription factor p53, involved in regulating cell cycle and apoptosis, is also activated during PC12 differentiation and contributes to each of these processes but the mechanisms are incompletely understood. NGF signaling stabilizes p53 protein expression, which enables its transcriptional regulation of target genes, including the newly identified target, wnt7b, a member of the wnt family of secreted morphogens. We tested the hypothesis that wnt7b expression is a factor in NGF-dependent neurite outgrowth of differentiating PC12 cells. Wnt7b transcript and protein levels are increased following NGF treatment in a p53-dependent manner, as demonstrated by a reduction in wnt7b protein levels following stable shRNA-mediated silencing of p53. In addition, overexpressed human tp53 was capable of inducing marked wnt7b expression in neuronal PC12 cells but tp53 overexpression did not elevate wnt7b levels in several tested human tumor cell lines. Ectopic wnt7b overexpression was sufficient to rescue neurite outgrowth in NGF-treated p53-silenced PC12 cells, which could be blocked by c-Jun N-terminal kinase (JNK) inhibition with SP600125 and did not involve beta-catenin nuclear translocation. Addition of sFRP1 to differentiation medium inhibited wnt7b-dependent phosphorylation of JNK, demonstrating that wnt7b is secreted and signals through a JNK-dependent mechanism in PC12 cells. We further identify an NGF-inducible subset of wnt receptors that likely supports wnt7b-mediated neurite extension in PC12 cells. In conclusion, wnt7b is a novel p53-regulated neuritogenic factor in PC12 cells that in conjunction with NGF-regulated Fzd expression is involved in p53-dependent neurite outgrowth through noncanonical JNK signaling.