Function and Regulation of TSPAN2 in Vascular Disease

TSPAN2在血管疾病中的功能和调控

• 批准号: 10083017
• 负责人: Xiaochun Long
• 金额: $ 52.22万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-08 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10083017
• 关键词: Adult; Allografting; Apolipoprotein E; Arteriovenous fistula; Atherosclerosis; Attenuated; Blood Vessels; CD44 gene; CRISPR/Cas technology; Cell Differentiation process; Cell membrane; Cell surface; Data; Dependence; Development; Disease; Docking; Electrophoretic Mobility Shift Assay; Endocytosis; Epigenetic Process; Exhibits; Failure; Gene Expression; Genes; Genetic Transcription; Goals; High Fat Diet; Homeostasis; Human; Impairment; In Vitro; Injury; Integrin Binding; Integrin alpha3beta1; Integrins; Knockout Mice; Knowledge; Laminin; Lead; Lesion; Link; Luciferases; Maintenance; Medial; Mediator of activation protein; Membrane; Modeling; Molecular; Mus; Mutant Strains Mice; Operative Surgical Procedures; Pathologic; Pathology; Pathway interactions; Phenotype; Proliferating; Proteins; RNA; Receptor Protein-Tyrosine Kinases; Regulation; Role; Sampling; Serum Response Factor; Signal Transduction; Smooth Muscle Myocytes; Stimulus; Testing; Therapeutic; Translating; Triad Acrylic Resin; Vascular Diseases; Vascular Smooth Muscle; blood pressure regulation; cell motility; effective therapy; genome editing; genome wide association study; improved; in vitro Assay; in vivo; loss of function; member; mouse model; myocardin; neointima formation; novel; programs; protein activation; protein degradation; response; restenosis; vascular abnormality; vascular smooth muscle cell proliferation

PROJECT SUMMARY Differentiated vascular smooth muscle cells (VSMCs) are genetically programmed to proliferate and migrate at low rate while carrying out contractility for adult vessel homeostasis. VSMCs are not terminally differentiated and can undergo phenotypic switching to a dedifferentiated synthetic mode in response to various pathophysiological stimuli, contributing to diverse vascular diseases such as atherosclerosis and restenosis. Due to cell surface accessibility, cell membrane regulators required for VSMC differentiation may hold attractive therapeutic potential. However, there is a paucity of information on key transmembrane regulators governing VSMC differentiation. TSPANs possess a unique signaling platform, termed as tetraspanin-enriched microdomains (TEMs) organized in cis with TSPANs and other transmembrane partners including receptor tyrosine kinases and integrin(s). TSPANs serve as membrane “docking” molecules, exerting critical roles in diverse signal cascades. Recent genome wide association studies have specifically linked TSPAN2 to atherosclerosis and blood pressure control. Inspired by these findings, we interrogated data from multiple RNA screenings and consistently found that TSPAN2 is the sole TSPANs member significantly decreased during VSMC phenotypic modulation and in diseased vessels. TSPAN2 is enriched in VSMCs and exhibits serum response factor (SRF) /Myocardin (MYOCD)-dependent expression in vitro. New functional data shows that depletion of TSPAN2 in VSMCs attenuates contractile gene expression, while promoting VSMC proliferation and migration; forced expression of TSPAN2 blocks neointima formation in a balloon injury model. TSPAN2 interacts with and promotes degradation of CD44 protein, a prominent pathological mediator in vascular disease. In addition, TSPAN2 interacts with the laminin-binding integrin α3β1, and β1 integrins are essential to the maintenance of the VSMC contractile phenotype. These preliminary findings support a novel hypothesis that TSPAN2 stabilizes the VSMC contractile phenotype and suppresses vascular pathology via two distinct pathways: by promoting degradation of CD44 protein which inactivates the synthetic VSMC phenotype and interaction with integrin α3β1 which facilitates the contractile phenotype. Three specific aims are proposed to test this hypothesis. Aim 1 will determine functions of TSPAN2 in arteriovenous fistula and atherosclerosis mouse models. Aim 2 will elucidate the mechanisms through which TSPAN2 stabilizes the VSMC contractile phenotype involving inactivation of the synthetic VSMC phenotype by promoting CD44 protein degradation and activation of the contractile phenotype by interacting with integrin α3β1. Aim 3 will elucidate the molecular basis for the impaired TSPAN2 gene expression under pathological vascular conditions via perturbation of SRF/MYOCD pathway. Successful completion of this proposal will reveal previously unknown molecular mechanisms governing VSMC phenotypic plasticity involving the actions of key transmembrane regulators, which will potentially lead to more effective therapies for vascular diseases.

项目总结 分化的血管平滑肌细胞(VSMCs)在基因上被编程为增殖和迁移 在进行成人血管内稳态收缩时的低率。VSMC不是终极分化的 并且可以响应于各种变化而进行表型切换到去分化合成模式 病理生理刺激，导致各种血管疾病，如动脉粥样硬化和再狭窄。 由于细胞表面的可及性，VSMC分化所需的细胞膜调节剂可能保持不变 诱人的治疗潜力。然而，关于关键的跨膜调节剂的信息很少。 管理VSMC分化。TSPAN具有独特的信号平台，称为富含TSPAN 与TSPAN和包括受体在内的其他跨膜伙伴在顺式结构中组织的微域(TEM) 酪氨酸激酶和整合素(S)。TSPAN是一种膜“对接”分子，在 不同的信号级联。最近的全基因组关联研究明确地将TSPAN2与 动脉粥样硬化和血压控制。受这些发现的启发，我们询问了来自多个RNA的数据 筛选并一致发现，TSPAN2是唯一的TSPANs成员，在 VSMC的表型调节和病变血管。TSPAN2富含血管平滑肌细胞并显示血清 反应因子(SRF)/Myocardin(MYOCD)依赖的体外表达。新的功能数据表明， VSMC中TSPAN2的缺失抑制收缩基因的表达，同时促进VSMC增殖 和迁移；在球囊损伤模型中，强制表达TSPAN2阻止了新生内膜的形成。TSPAN2 与CD44蛋白相互作用并促进其降解，CD44蛋白是血管中重要的病理介质 疾病。此外，TSPAN2与层粘连蛋白结合的整合素α3β1相互作用，而β1整合素是必不可少的 VSMC收缩表型的维持。这些初步发现支持一种新的假设。 TSPAN2通过两种不同的途径稳定VSMC的收缩表型并抑制血管病变 途径：通过促进CD44蛋白的降解，使合成的VSMC表型失活 与整合素α3β1相互作用，促进收缩表型。提出了三个具体目标来 检验这一假设。目的1确定TSPAN2在动静脉瘘和动脉粥样硬化中的作用 老鼠模型。目标2将阐明TSPAN2稳定VSMC收缩的机制 表型包括通过促进CD44蛋白降解和促进合成的VSMC表型失活 通过与整合素α3β1相互作用激活收缩表型。Aim 3将阐明分子 TSPAN2基因在病理性血管条件下表达受损的基础 SRF/MYOCD途径。这项计划的成功完成将揭示以前未知的分子 调控VSMC表型可塑性的机制涉及到关键的跨膜调节因子的作用， 这可能会带来更有效的血管疾病治疗方法。