Nuclear translocation of urokinase/nucleolin complexes

尿激酶/核仁素复合物的核转位

基本信息

批准号：
7319578
负责人：
Douglas Brock Cines
金额：
$ 22.94万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-07-01 至 2009-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7319578
关键词：
Address Aneurysm Atherosclerosis Binding Binding Sites Biological Biological Assay Biology Blood Vessels C-terminal Candidate Disease Gene Cardiovascular Diseases Cell Nucleus Cell Proliferation Cell surface Cells Chromatin Complement Complex Conserved Sequence Coronary Coronary artery DNA DNA Binding DNA Microarray Chip DNA Microarray format Data Electrophoretic Mobility Shift Assay Endothelial Cells Fibroblasts Gene Expression Gene Expression Regulation Genes Genetic Transcription Goals Growth Growth Factor Human Hypertension Immigration In Vitro Inflammation Knowledge Kringles Mammalian Cell Mediating Membrane Microarray Analysis Mus Neoplasm Metastasis Nuclear Nuclear Import Nuclear Protein Nuclear Proteins Nuclear Translocation Pathogenesis Pathologic Processes Pathway interactions Personal Satisfaction Phenotype Pilot Projects Plasmin Plasminogen Activator Plasminogen Activator Inhibitor 1 Polymerase Chain Reaction Precipitation Process Property Protease Domain Protein Microchips Protein Overexpression Proteins Proteolysis Reporter Reporting Reverse Transcriptase Polymerase Chain Reaction Screening procedure Serine Protease Signal Transduction Single-Stranded DNA Site-Directed Mutagenesis Smooth Muscle Myocytes Surface System Time Urokinase Vascular Diseases Vascular remodeling Western Blotting Wound Healing Yeasts angiogenesis atherogenesis base bone morphogenetic protein 6 cell motility ds-DNA fibulin 1 immunocytochemistry in vivo inhibitor/antagonist insight interest migration mouse Smc1l1 protein mouse Smc1l2 protein novel novel therapeutics nucleocytoplasmic transport nucleolin prevent promoter receptor response restenosis therapeutic target trafficking transcription factor tumor vascular smooth muscle cell proliferation yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): Vascular remodeling is an essential component in the pathogenesis of atherosclerosis and other prevalent cardiovascular disorders, such as aneurysm formation, restenosis, and hypertension. The involvement of plasminogen activator (PA)/plasmin system in atherogenesis is well established through multiple studies that demonstrate overexpression of uPA, tPA and their inhibitor PAI-1 in human athrosclerosis. uPA and its surface receptor (uPAR) are implicated in vascular SMC proliferation/migration and matrix synthesis, which is pivotal in the progression of atherosclerosis. Our studies are the first to indicate that scuPA is translocated to the nucleus of mammalian cells in vitro and in vivo and we have identified one critical intermediary in the process, nucleolin. The issue we propose to address in this application is to delineate how uPA functions within the nucleus with the long-term goal of relating nuclear translocation and gene transcription to uPA-mediated alterations in cell adhesivity, proliferation and migration in vascular remodeling and atherosclerosis. Using Affymetrix DNA microarray, we have identified at least 6 genes relevant to vascular remodeling that are induced by uPA in nucleolin-dependent manner. In Aim 1, we propose to validate these microarray findings in coronary arterial SMC using RT-PCR and western blotting to provide a biological underpinning for what follows. In Aim 2 we will follow results already obtained from transcription factor (TF) arrays and use a mammalian two-hybrid system to determine whether scuPA or nucleolin bind directly to these or other TFs or to novel partners that regulate nuclear transport or transcription. In Aim 3, we will explore an alternative and novel mechanism by asking whether uPA regulates transcription by binding directly to DNA and/or gene promoters. Together these studies will elucidate a new facet of uPA biology and should provide an opportunity to develop specific means to regulate uPA- mediated pathologic processes. This knowledge will help to identify novel therapeutic approaches to prevent cell invasion and growth in vascular remodeling. This project intends to investigate how urokinase-type plasminogen activator uPA) stimulates gene expression, proliferation and migration in vascular SMC. These studies will reveal a novel pathway and novel targets to interrupt pathogenic intimal SMC migration in atherosclerosis and other common human vascular disorders as well as in progression of many human tumors.

描述（由申请人提供）：血管重塑是动脉粥样硬化和其他常见心血管疾病（例如动脉瘤形成、再狭窄和高血压）发病机制的重要组成部分。多项研究证实了纤溶酶原激活剂 (PA)/纤溶酶系统参与动脉粥样硬化形成，这些研究证明了人类动脉粥样硬化中 uPA、tPA 及其抑制剂 PAI-1 的过度表达。 uPA 及其表面受体 (uPAR) 参与血管 SMC 增殖/迁移和基质合成，这在动脉粥样硬化的进展中至关重要。我们的研究首次表明 scuPA 在体外和体内易位至哺乳动物细胞的细胞核，并且我们已经确定了该过程中的一个关键中间体：核仁素。我们建议在本申请中解决的问题是描述uPA如何在细胞核内发挥作用，长期目标是将核易位和基因转录与血管重塑和动脉粥样硬化中uPA介导的细胞粘附性、增殖和迁移的改变联系起来。使用 Affymetrix DNA 微阵列，我们鉴定了至少 6 个与血管重塑相关的基因，这些基因是由 uPA 以核仁素依赖性方式诱导的。在目标 1 中，我们建议使用 RT-PCR 和蛋白质印迹法验证冠状动脉 SMC 中的这些微阵列结果，为后续研究提供生物学基础。在目标 2 中，我们将跟踪从转录因子 (TF) 阵列获得的结果，并使用哺乳动物双杂交系统来确定 scuPA 或核仁素是否直接与这些或其他 TF 或调节核运输或转录的新伙伴结合。在目标 3 中，我们将通过询问 uPA 是否通过直接与 DNA 和/或基因启动子结合来调节转录来探索一种替代的新颖机制。这些研究将共同阐明 uPA 生物学的一个新方面，并为开发调节 uPA 介导的病理过程的特定方法提供机会。这些知识将有助于确定新的治疗方法，以防止血管重塑中的细胞侵袭和生长。该项目旨在研究尿激酶型纤溶酶原激活剂（uPA）如何刺激血管平滑肌细胞中的基因表达、增殖和迁移。这些研究将揭示一种新的途径和新的靶标，以中断动脉粥样硬化和其他常见人类血管疾病以及许多人类肿瘤进展中的致病性内膜 SMC 迁移。