Yoon J K, Lau L F
Department of Genetics, University of Illinois College of Medicine, Chicago 60612.
Mol Cell Biol. 1994 Dec;14(12):7731-43. doi: 10.1128/mcb.14.12.7731-7743.1994.
nur77, an immediate-early gene that encodes an orphan nuclear receptor, is rapidly and transiently induced by nerve growth factor (NGF) stimulation or membrane depolarization in the rat pheochromocytoma-derived cell line PC12. The Nur77 protein can act as a potent transcription activator and may function to regulate the expression of downstream genes in response to extracellular stimuli. We show here that activation of nur77 by NGF treatment and membrane depolarization is signalled through distinct pathways. These distinct signals appear to converge on the same transcription factors acting on the same promoter elements. We show that nur77 activation by both processes requires two cis-acting AP1-like elements, NAP1 and NAP2, which contain the core sequence TGCGTCA centered at 67 and 38 nucleotides upstream of the transcription start site. The NAP elements can confer inducibility by NGF and membrane depolarization on an otherwise unresponsive heterologous promoter. We identified JunD as a key mediator of nur77 activation by reason of the following observations. (i) JunD, but not CREB or other members of the Fos/Jun family, is a component of NAP binding activity in PC12 cell nuclear extracts. (ii) JunD, but not other Fos/Jun family members, specifically transactivates the nur77 promoter through the NAP elements (iii) A dominant-negative mutant of JunD effectively abolishes the activation of nur77 by either NGF treatment or membrane depolarization. These data draw a contrast between the regulation of nur77 with that of c-fos, in which the sequence requirements for activation by NGF treatment and membrane depolarization appear separable, and CREB appears to play a role in activation by both NGF and membrane depolarization.
Nur77是一种立即早期基因，编码一种孤儿核受体，在大鼠嗜铬细胞瘤衍生的PC12细胞系中，它可被神经生长因子（NGF）刺激或膜去极化迅速且短暂地诱导。Nur77蛋白可作为一种强效转录激活因子，可能在响应细胞外刺激时调节下游基因的表达。我们在此表明，NGF处理和膜去极化对Nur77的激活是通过不同途径发出信号的。这些不同的信号似乎汇聚于作用于相同启动子元件的相同转录因子。我们表明，这两种过程对Nur77的激活都需要两个顺式作用的AP1样元件，即NAP1和NAP2，它们包含位于转录起始位点上游67和38个核苷酸处的核心序列TGCGTCA。NAP元件可赋予NGF和膜去极化对原本无反应的异源启动子的诱导性。基于以下观察结果，我们确定JunD是Nur77激活的关键介质。（i）JunD而非CREB或Fos/Jun家族的其他成员，是PC12细胞核提取物中NAP结合活性的一个组成部分。（ii）JunD而非其他Fos/Jun家族成员，通过NAP元件特异性地反式激活Nur77启动子。（iii）JunD的显性负性突变体有效地消除了NGF处理或膜去极化对Nur77的激活。这些数据将Nur77的调节与c-fos的调节进行了对比，在c-fos中，NGF处理和膜去极化激活的序列要求似乎是可分离的，而CREB似乎在NGF和膜去极化的激活中都发挥作用。
