CONTROL OF MONOCYTE CHEMOTACTIC FACTOR EXPRESSION

这个项目的长期目标是了解细胞内的 细胞因子和激素调节转录的机制 否则就是静止的基因。血小板对成纤维细胞的刺激作用 衍生生长因子(PDGF)导致血管内皮细胞 我是吉恩。乙脑编码一种单核细胞的小鼠同源蛋白 趋化蛋白-I(MCP-1)是一个不断增长的超家族成员 相关的可诱导细胞因子。JE/MCP-1已被确定为主要 来自平滑肌细胞的单核细胞特异性趋化因子和各种 肿瘤细胞系，并可能在肿瘤的发病机制中发挥重要作用。 动脉粥样硬化，在伤口愈合和炎症过程中。强有力的 抗炎糖皮质激素抑制PDGF诱导的乙脑 具有与其药理一致的药效等级顺序的基因 作为一种消炎剂。DNA的缺失和突变 乙脑基因两侧或内部的序列将被用来识别关键字 PDGF诱导转录的调控序列 糖皮质激素的抑制作用。结合转录因子 将通过足迹和凝胶来表征这些序列 发育迟缓分析。这些因子的大规模提纯将是 他们的基因将被克隆。实验旨在 糖皮质激素介导的四种模型机制辨析 转录抑制被提出。为了进一步深入研究 乙脑转录活性的正负调节， 提出了以下具体目标： 特异性目标1：乙脑启动子缺失突变体的构建与分析 识别转录增强子区(PDGF反应元件) 负责PDGF诱导的基因表达。 具体目标2：定位和确定负责 糖皮质激素介导的转录抑制。 具体目标3：鉴定和鉴定转录因子 识别和结合PDGF反应元件和糖皮质激素介导的 转录抑制物区。 具体目标4：将亲缘关系、数目和可能的亲属联系起来 转录因子DNA与转录因子水平结合的位置效应 PDGF诱导JE表达和糖皮质激素介导的抑制。 特定目标5：纯化和克隆转录因子 参与PDGF对JE基因的正向调控 糖皮质激素的负性调节。

The long term goal of this project is to understand the intracellular mechanism by which cytokines and hormones regulate the transcription of otherwise quiescent genes. Stimulation of fibroblasts with Platelet- derived growth factor (PDGF) results in the increased transcription of the JE gene. JE encodes a protein in which is the murine homolog of Monocyte Chemotactic Protein-I (MCP-1) and is a member of a growing superfamily of related inducible cytokines. JE/MCP-1 has been identified as the major monocyte specific chemotactic factor from smooth muscle cells and various tumor cell lines, and may play an important role in the pathogenesis of atherosclerosis, in wound healing and in the inflammatory process. Potent anti-inflammatory glucocorticoids inhibits the PDGF induction of the JE gene with the same rank order of potency consistent with its pharmacology as an anti-inflammatory agent. Deletions and mutations of the DNA sequences flanking or within the JE gene will be made to identify key regulatory sequences responsible for the transcriptional induction by PDGF and the repression by glucocorticoids. Transcriptional factors which bind to these sequences will be characterized by footprinting and gel retardation assays. Large scale purification of these factors will be attempted and their genes will be cloned. Experiments designed to distinguish between four model mechanisms of glucocorticoid mediated transcriptional repression are proposed. To further the study of the positive and negative regulation of JE transcriptional activity, the following specific aims are proposed: Specific Aim 1: Construction and analysis of JE promotor deletion mutants to identify transcriptional enhancer regions (PDGF response elements) which are responsible for PDGF induced gene expression. Specific Aim 2: Locate and identify DNA repressor regions responsible for the glucocorticoid mediated transcriptional repression. Specific Aim 3: To identify and characterize transcription factors which recognize and bind to PDGF response elements and to glucocorticoid mediated transcriptional repressor regions. Specific Aim 4: To correlate the affinity, number and possible relative positional effects of transcription factor DNA binding with the level of PDGF induced JE expression and glucocorticoid mediated repression. Specific Aim 5: To purify and clone transcription factors which are involved in the positive regulation of the JE gene by PDGF and in the negative regulation by glucocorticoids.