The STIM/Orai interface and novel tools to control vascular smooth muscle phenotype

STIM/Orai 接口和控制血管平滑肌表型的新工具

• 批准号: 10431766
• 负责人: James Henry Baraniak
• 金额: $ 1.78万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10431766
• 关键词: Address; Affinity; Amaze; Apoptosis; Atherosclerosis; Binding; Binding Sites; Biological Assay; Blood Vessels; Blood flow; C-terminal; Calcium; Calcium Channel; Calcium Signaling; Cardiovascular Diseases; Cardiovascular system; Cell membrane; Cell physiology; Cells; Clinical; Complex; Couples; Coupling; Development; Disease; Electrophysiology (science); Endoplasmic Reticulum; Event; Fluorescence Recovery After Photobleaching; Fluorescence Resonance Energy Transfer; Fura-2; Gene Expression; Generations; Goals; Growth; Hybrids; IL2 gene; Image; Impairment; Inflammation; Injury; Knowledge; Lead; Luciferases; Measures; Mechanics; Mediating; Modification; Molecular Conformation; Nature; Nuclear; Pathogenicity; Pathologic; Peptides; Phenotype; Physiological; Plasmids; Process; Property; Protein Family; Proteins; Quantitative Reverse Transcriptase PCR; Research; Resolution; STIM1 gene; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Smooth Muscle Myocytes; Structure; Systemic hypertension; T-Cell Activation; T-Lymphocyte; Testing; Therapeutic; Training; Vascular Proliferation; Vascular Smooth Muscle; Work; base; cell growth; cell motility; cell type; crosslink; density; dimer; experimental study; flexibility; in vivo; migration; neointima formation; novel; novel therapeutics; patch clamp; prevent; receptor; receptor-mediated signaling; response to injury; restenosis; tool; transcription factor; vascular smooth muscle cell migration; vascular smooth muscle cell proliferation; voltage; wound; wound healing; wound injury; wound response

PROJECT ABSTRACT Vascular smooth muscle cells (VSMCs) are crucial in maintaining the integrity of vessel walls, vascular tone, and contraction. They have the amazing potential to dedifferentiate upon mechanical injury or inflammation from a quiescent state to a proliferative (synthetic) state. This high degree of VSMC plasticity is crucial in wound response and healing, but also predisposes the cell to adverse formation of neointima near the vascular lumen. Neointimal formation tightens the arterial wall, restricting blood flow and is the pathogenic base for occlusive cardiovascular diseases such as atherosclerosis, restenosis, and systemic hypertension. Therapies to address neointimal formation are scarce and non-selective. Understanding and targeting mechanisms utilized by synthetic VSMCs is of upmost clinical importance. What has been observed in the transition between quiescent and synthetic VSMCs, is a phenotypic modulation of proteins involved in calcium signaling. Calcium is a vital second messenger involved in nearly every cellular process from proliferation to apoptosis. As VSMCs change from their contractile quiescent state to a proliferative state, they rely less on voltage-gated calcium channels and depend more on the Orai1 calcium channel. Orai1 is essential in the calcium homeostatic process known as store operated calcium entry (SOCE). Calcium stores within the endoplasmic reticulum (ER) are sensed by a family of proteins known as STIM. Upon store depletion, STIM proteins oligomerize, undergo conformational change, and migrate from ER to ER-plasma membrane (PM) junctions. Unfolding of the STIM reveals a powerful interacting site known as the STIM-Orai activating region (SOAR), which is able to bind and activate the highly calcium selective PM Orai channels. Elucidation and exploitation of the STIM-Orai binding interface presents an exciting new therapeutic avenue for cardiovascular disease. How Orai couples with STIM is a topic under much speculation. Previous studies propose that either two Orai cytosolic regions are involved, known as the N-terminus and transmembrane extension (TM4ext), or that adjacent TM4exts are required to form a dimerized binding pocket. In order to solve the mystery of the STIM- Orai coupling interface, I developed a novel set of PM tethering peptides that can exclusively interrogate the TM4ext of any of the three major mammalian Orai subtypes, Orai1-3. My recent work presents an exciting new paradigm for STIM-Orai interacting, that the TM4ext is necessary and sufficient in acting as a sole binding region. Furthermore, I found out that the Orai3 TM4ext ahs tremendous binding interaction with STIM over the more ubiquitous Orai1. This F31 proposal builds upon these exciting developments and represents a huge integral part for the training of James Baraniak. Continual work on this proposal will interrogate how STIM-Orai interacts using these novel peptides, and how they may be used as a powerful tool in controlling VSMC proliferation and migration.

项目摘要 血管平滑肌细胞（VSMC）在维持血管壁的完整性、血管内皮细胞的增殖和分化中起着至关重要的作用。 音调和收缩。它们具有在机械损伤或炎症时去分化的惊人潜力 从静止状态到增殖（合成）状态。这种高度的VSMC可塑性在创伤中至关重要， 这不仅影响了细胞的反应和愈合，而且还使细胞易于在血管腔附近不利地形成新生内膜。 新生内膜形成使动脉壁变紧，限制血流，并且是闭塞性血管病变的致病基础。 心血管疾病，如动脉粥样硬化、再狭窄和全身性高血压。治疗方法 新生内膜形成是罕见和非选择性的。了解和确定目标的机制， 合成的VSMC具有最重要的临床意义。 在静止和合成VSMC之间的过渡中观察到的是一种表型， 调节参与钙信号传导的蛋白质。钙是一种重要的第二信使， 从增殖到凋亡的所有细胞过程。当血管平滑肌细胞从收缩的静止状态 到增殖状态，它们较少依赖于电压门控钙通道，而更多地依赖于Orai 1钙通道。 频道Orai 1在钙稳态过程中是必不可少的，称为钙库操纵的钙进入（SOCE）。 内质网（ER）内的钙储存由一个称为STIM的蛋白质家族感知。后 储存耗尽，STIM蛋白寡聚化，经历构象变化，并从ER迁移到ER-血浆 膜（PM）连接。STIM的展开揭示了一个强大的相互作用位点，称为STIM-Orai 激活区（SOAR），其能够结合并激活高钙选择性PM奥赖通道。 STIM-Orai结合界面的阐明和开发提出了一种令人兴奋的新治疗方法， 心血管疾病的途径。 奥赖如何与STIM配对是一个备受猜测的话题。先前的研究表明， 涉及奥赖胞质区域，称为N-末端和跨膜延伸（TM 4 ext），或者 需要相邻的TM 4 ext来形成二聚结合口袋。为了解开“性刺激”之谜- 奥赖耦合接口，我开发了一套新的PM拴系肽，可以专门询问 三种主要的哺乳动物奥赖亚型之一的TM 4 ext，奥莱1 -3。我最近的工作展示了一个令人兴奋的新的 对于STIM-Orai相互作用的范例，TM 4 ext在充当唯一结合区域方面是必要且充分的。 此外，我发现Orai 3 TM 4 ext与STIM的结合相互作用超过了 无处不在的Orai 1. F31提案建立在这些令人兴奋的发展基础上，代表了一个巨大的组成部分， 詹姆斯·巴拉尼亚克的训练关于该提案的持续工作将询问STIM-Orai如何使用这些 新的肽，以及它们如何可以用作控制VSMC增殖和迁移的有力工具。