DYNAMICS OF C-FOS PROTEIN INTERACTIONS

我们的实验室最近发现，c-fos的诱导表达 基因是一种细胞对多种跨膜信号的反应 药物包括生长因子、佛波酯和神经递质。我们 建议研究Fos蛋白的性质及其诱导作用。 具体来说，我们建议： 1.测定Fos蛋白合成的时间进程 生长后Fos与第二种细胞蛋白P39的相关性 3T3细胞中的因子诱导。 2.测定FOS蛋白与FOS蛋白复合物的相对分子质量 第39页。从化学计量学的变化中，我们将推断出 FOS-P39复合体形成。 3.诱变FOS蛋白。我们将确定结合部位 并将其与改变FOS蛋白的突变位点进行比较 转化能力、核定位、翻译后修饰 和表达的自动调节的可能性。这将开发一种 FOS蛋白的功能拓扑。 4.分离P39蛋白与FOS的复合体，纯化P39蛋白 目的制备研究P39表达和结构的试剂。 5.检测FOS蛋白在细胞内的负性调节作用 FOS启动子。 6.检测其他生长因子诱导的FOS早期反应基因 使用反义技术的转录抑制活性。 7.鉴定与FOS调控序列相互作用的蛋白质 新型DNA标记转移分析。

Our laboratory has recently shown that induction of expression of the c-fos gene is a cellular response to a wide range of transmembrane signaling agents including growth factors, phorbol esters, and neurotransmitters. We proposed to study properties of the fos protein and its induction. Specifically we propose: 1. To determine the time course of synthesis of fos protein and the association of fos with a second cellular protein, p39 following growth factor induction in 3T3 cells. 2. To determine the molecular weight of the complex of fos protein with p39. From changes in this stoichiometry we will deduce the dynamics of fos-p39 complex formation. 3. To mutagenize the fos protein. We will determine the binding site on fos for p39 and compare this with sites of mutation which alter fos protein transforming ability, nuclear localization, post-translational modification and potential for autoregulation of expression. This will develop a functional topology for the fos protein. 4. To purify the p39 protein through isolation of its complex with fos with the objective of preparing reagents to study p39 expression and structure. 5. To assay the fos protein for a negative regulatory role acting at the fos promoter. 6. To assay other growth factor induced early response genes for fos transcription repressor activity using the antisense technique. 7. To identify proteins interacting with fos regulatory sequences using a novel DNA label transfer assay.