DYNAMICS OF C-FOS PROTEIN INTERACTIONS

fos蛋白是一种核DNA结合蛋白， 调节因子，并在广泛的细胞类型中由许多不同的 跨膜信号，包括生长因子刺激。 近日 已经表明fos与jun蛋白质形成特异性复合物 通过含有以下七肽重复序列的α螺旋区的相互作用， 亮氨酸（亮氨酸拉链）。 拉链式相互作用使碱性氨基 两种蛋白质的酸性基序形成序列特异性DNA结合 域 我们建议确定fos的结构特征， 特异性异源二聚化与jun。这些可能存在于 α螺旋，或其他地方，并将分析的方法，网站 定向诱变和网织红细胞裂解物翻译的体外测定 产品. 我们还表明，当神经生长因子（NGF） 刺激模型PC 12嗜铬细胞瘤细胞分化， 神经元通路，诱导Fos。 fos转录的高峰是在 神经生长因子 在fos诱导后，其他表达神经元特异性的基因， 功能也被表达出来。 酪氨酸羟化酶（TH） 基因，在NGF处理后1小时转录诱导。 的TH 启动子具有结合真正的fos-jun复合物的DNA元件， 结合由NGF在1-4周期内诱导的体内PC 12产物 hr处理后。 这与TH转录 受到积极和消极的调节。 我们将分析诱导的PC 12 可能调节TH表达的因子。 这将包括 鉴定与TH基因启动子相互作用的因子， 分析fos的特殊作用，并寻找新的， Fos和Jun家族的神经元特异性成员。 因此，具体 该提案的目标是：目标1。 确定的结构特征 Fos赋予Fos和Jun异二聚化的特异性 proteins. 目标二。 确定立即早期反应的作用 基因（包括fos家族成员）在转录调控 PC 12细胞延迟早期基因。

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.