DYNAMICS OF C-FOS PROTEIN INTERACTIONS

Fos蛋白是一种核DNA结合蛋白，它是一种转录 调节因子，并在多种细胞类型中被多种不同的 跨膜信号，包括生长因子刺激。最近，它 已经证明，FOS与Jun蛋白形成一种特定的复合体 通过包含七肽重复的α螺旋区域的相互作用 亮氨酸(亮氨酸拉链)拉链相互作用与碱性氨基并列 形成序列特异性DNA结合的两种蛋白质的酸性基序 域。我们建议确定FOS的结构特征，这些特征给出了 与Jun的异二聚化的特异性。它们可以驻留在 α螺旋，或其他地方，并将通过现场的方法进行分析 网织红细胞裂解物翻译的定向诱变及体外检测 产品。我们还表明，当神经生长因子(NGF) 刺激PC12嗜铬细胞瘤模型细胞向下分化 神经通路，FOS被诱导。Fos转录高峰期为30min。开机自检 NGF。在Fos诱导后，表达神经元特异性的其他基因 功能也被表达出来。其中之一是酪氨酸羟基酶(TH) 基因，转录诱导1小时。在NGF治疗后。这是 启动子具有与正品FOS-JUN复合体结合的DNA元件，还 结合体内由NGF诱导的PC12产物1-4 HR治疗后。这与转录的时间间隔相同 受到积极和消极的监管。我们将分析诱导的PC12 寻找可能调节TH表达的因素。这将包括 与TH基因启动子相互作用的因子)S)， 分析FOS的特殊作用，并寻找新的、 Fos和Jun家族中神经元特有的成员。因此，具体的 建议的目的是：目的1.确定结构特征 赋予FOS和JUN异二聚化专一性的FOS 蛋白质。目标2.确定即时早期反应的作用 转录调控基因(包括FOS家族成员) 延迟了PC12细胞中的早期基因。

The fos protein is a nuclear, DNA binding protein which is a transcription regulator and is induced in a wide range of cell types by many different transmembrane signals, including growth factor stimulation. Recently, it has been shown that fos makes a specific complex with the jun protein through interactions of alpha helical regions containing heptad repeats of leucines (leucine zipper). The zipper interaction juxtaposes basic amino acid motifs of both proteins which form a sequence specific DNA binding domain. We propose to determine the structural features of fos which give specificity for heterodimerization with jun. These could reside within the alpha helix, or elsewhere, and will be analyzed by the method of site directed mutagenesis and in vitro assay of reticulocyte lysate translation products. We have also shown that when nerve growth factor (NGF) stimulates model PC12 pheochromocytoma cells to differentiate down a neuronal pathway, fos is induced. Peak fos transcription is 30 min. post NGF. Following fos induction, other genes which express neuronal specific functions are also expressed. One of these, the tyrosien hydroxylase (TH) gene, is induced transcriptionally 1 hr. following NGF treatment. The TH promoter has a DNA element which binds authentic fos-jun complex and also binds an in vivo PC12 product which is induced by NGF over the period 1-4 hr post treatment. This is the same interval over which TH transcription is positively and negatively regulated. We will analyze the induced PC12 for factors which may regulate TH expression. This will include identification of the factor)s) which interact with the TH gene promoter, analysis of the particular role of fos, and a search for new, neuronal-specific members of the fos and jun families. Thus, the Specific Aims of the proposal are: Aim 1. To determine the structural features of fos which confer specificity of heterodimerization on the fos and jun proteins. Aim 2. To determine the role of the immediate early response genes (including the fos family members) in regulation of transcription of delayed early genes in PC12 cells.