Brynczka Christopher, Merrick B Alex
National Center for Toxicogenomics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709, USA.
Differentiation. 2008 Sep;76(7):795-808. doi: 10.1111/j.1432-0436.2007.00261.x. Epub 2008 Jan 3.
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.
神经生长因子（NGF）诱导PC12细胞分化的特征在于信号转导途径的变化，这种变化会导致细胞生长停滞和神经突延伸。转录因子p53参与调节细胞周期和细胞凋亡，在PC12细胞分化过程中也被激活，并对上述每个过程都有作用，但其机制尚不完全清楚。NGF信号传导可稳定p53蛋白表达，从而使其能够对靶基因进行转录调控，这些靶基因包括新发现的靶标wnt7b，它是分泌型形态发生素wnt家族的成员。我们验证了一个假设，即wnt7b表达是PC12细胞分化过程中NGF依赖性神经突生长的一个因素。如稳定的shRNA介导的p53沉默后wnt7b蛋白水平降低所示，NGF处理后，wnt7b转录本和蛋白水平以p53依赖性方式增加。此外，过表达的人tp53能够在神经元PC12细胞中诱导显著的wnt7b表达，但tp53过表达并未提高几种测试的人肿瘤细胞系中的wnt7b水平。异位过表达wnt7b足以挽救NGF处理的p53沉默的PC12细胞中的神经突生长，这可被SP600125抑制c-Jun N端激酶（JNK）所阻断，且不涉及β-连环蛋白核转位。向分化培养基中添加sFRP1可抑制wnt7b依赖性的JNK磷酸化，表明wnt7b被分泌并通过PC12细胞中的JNK依赖性机制发出信号。我们进一步鉴定了一个NGF诱导的wnt受体亚群，其可能支持PC12细胞中wnt7b介导的神经突延伸。总之，wnt7b是PC12细胞中一种新的p53调节的神经生成因子，它与NGF调节的Fzd表达一起，通过非经典JNK信号传导参与p53依赖性神经突生长。
