NUCLEAR INTEGRATION OF ENDOTHELIAL SIGNALS

内皮信号的核整合

• 批准号: 6469266
• 负责人: Tucker O Collins
• 金额: $ 6.87万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6469266
• 关键词: cAMP response element binding protein; calcium binding protein; gel mobility shift assay; gene induction /repression; genetic promoter element; genetic regulation; genetic transcription; genetically modified animals; human tissue; immunocytochemistry; intermolecular interaction; laboratory mouse; leukocyte adhesion molecules; nuclear factor kappa beta; phosphoprotein phosphatase; selectins; tissue /cell culture; transcription factor; transfection; vascular endothelium

Expression of endothelial-leukocyte adhesion molecule is dramatically induced by inflammatory cytokines. This increased expression occurs at the transcriptional level and requires activation of the nuclear factor-kappaB (NF-kB) system. Recent studies indicate that the p65 component of NF-kB, like many signal dependent transcriptional activators, interacts with the transcriptional co-activator CREB-binding protein (CBP). In this continuing project, we will investigate whether CBP is part of a novel level of nuclear regulation linking diverse signaling systems in endothelial cells and test the hypothesis that competition for limiting amounts of CBP facilitates the activation of some signal dependent genes at the expense of others. In the renewal period we propose to determine if this regulatory system is relevant to the process of endothelial activation during inflammatory responses. In Specific Aim #1, the antagonistic interactions between p65 and other signal-dependent transcription factors occur because for limiting amounts of the common transcriptional co-activator, CBP. These studies will examine the levels of CBP expression in quiescent and cytokine-activated cultured endothelial cells, determine if levels of the co-activator limit NF-kappaB dependent gene expression, and will explore the molecular basis of the antagonistic interaction. In Specific Aim #2, the pathophysiologic consequences of CBP over- expression in endothelial cells will be examined in a transgenic model of endothelial activation. In this defined genetic context, we will determine if CBP over expression increases induction of NF-kappaB dependent genes and diminishes the negative regulatory effects of the activated nuclear receptors on the induction of kappaB-dependent genes. Like the induction process, decreased expression of the endothelial adhesion molecules following cytokine exposure is an active process that occurs at the transcriptional level. This level of negative regulation my be key in preventing inappropriate or prolonged expression of some adhesion molecule genes. In Specific Aim #3, the mechanisms responsible for the post-induction transcriptional repression of E-selectin will continue to be investigated. These studies will determine the role of the dual specificity phosphatases in limiting the expression of this adhesion molecule gene using tools developed in this project, as well as unique cellular and animal reagents developed by collaborators. Collectively, the findings from these studies should provide insights into two new endothelial regulatory systems: first, they should indicate if CBP serves an integration function for interactions between p65 and other classes of CBP dependent transcription factors during cytokine-induced gene expression; and second, they should define the mechanisms which tightly control the post-induction transcriptional repression of the E- selectin gene.

内皮细胞-白细胞黏附分子的表达显著增加 由炎性细胞因子诱导。这种增加的表达发生在 转录水平，需要核因子-kappaB的激活 (核因子-kB)系统。最近的研究表明，核因子-kB的p65组分， 像许多依赖信号的转录激活因子一样，它与 转录共激活因子CREB结合蛋白(CBP)。在这 继续项目，我们将调查CBP是否是小说的一部分 连接不同信号系统的核监管水平 并测试限制竞争的假说 大量的CBP促进一些信号依赖基因的激活 以牺牲他人为代价。在续约期内，我们建议确定是否 这一调节系统与内皮细胞的过程有关。 炎症反应过程中的激活。 在特定目标1中，p65和其他基因之间的拮抗相互作用 信号依赖的转录因子的产生是因为 共同的转录辅助激活因子CBP。这些研究将 检测静止期和细胞因子激活状态下CBP的表达水平 培养的内皮细胞，确定共激活因子的水平是否限制 核因子-kappaB依赖的基因表达，并将探讨其分子基础 对抗性的相互作用。 在具体目标#2中，CBP的病理生理后果超过- 将在转基因模型中检测血管内皮细胞的表达 血管内皮细胞激活。在这个明确的遗传背景下，我们将确定 如果CBP过度表达增加了对核因子-kappaB依赖基因的诱导 并减少了激活的核的负面调节效应 受体对kappaB依赖基因的诱导。 像诱导过程一样，内皮细胞的表达减少 细胞因子暴露后的黏附分子是一个活跃的过程， 发生在转录水平。这种程度的负面监管可能会 在防止某些不适当或长时间表达方面起到关键作用 黏附分子基因。 在具体目标3中，负责后诱导的机制 E-选择素的转录抑制作用将继续被研究。 这些研究将确定双特异性磷酸酶的作用。 使用工具限制该黏附分子基因的表达 在这个项目中开发的，以及独特的细胞和动物试剂 由合作者开发。 总体而言，这些研究的结果应该为以下方面提供见解 两个新的内皮调节系统：首先，它们应该表明CBP 为P65和其他之间的交互提供集成功能 细胞因子诱导过程中CBP依赖转录因子的分类 基因表达；第二，他们应该定义 严格控制诱导后对E-的转录抑制 选择素基因。