The NHERF1-STIM1-Orai1 axis regulates MRGPRX2 responses in mast cells

NHERF1-STIM1-Orai1 轴调节肥大细胞中的 MRGPRX2 反应

• 批准号: 10538916
• 负责人: Hariharan Subramanian
• 金额: $ 20万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10538916
• 关键词: Ablation; Adaptor Signaling Protein; Affect; Agent 48-80; Agonist; Allergic; Allergic Disease; Allergic Reaction; Anaphylaxis; Asthma; Biological Assay; CD34 gene; Cell Compartmentation; Cell physiology; Cells; Cellular biology; Chronic; Clinical; Couples; Coupling; Data; Development; Disease; Edema; Future; G-Protein-Coupled Receptors; Genetic; Goals; Grant; Human; Immune; In Vitro; Inflammation; Inflammatory; Intervention; Mediating; Molecular; Mus; National Heart, Lung, and Blood Institute; National Institute of Allergy and Infectious Disease; Orthologous Gene; Pathogenesis; Pathology; Pathway interactions; Pharmaceutical Preparations; Pharmacology; Play; Proteins; Reaction; Research; Role; Rosacea; STIM1 gene; Signal Transduction; Testing; TimeLine; Tissues; Umbilical Cord Blood; United States Food and Drug Administration; United States National Institutes of Health; Urticaria; allergic response; cathelicidin; cell type; chronic inflammatory disease; clinical application; cytokine; drug development; experimental study; in vivo; in vivo Model; insight; knock-down; mast cell; mouse model; mutant; novel; novel therapeutics; receptor; response; sensor; sodium-hydrogen exchanger regulatory factor

PROJECT SUMMARY Mast cells are tissue resident innate immune cells that have been best characterized for their role in mediating allergic diseases. A recent development in mast cell research has been the identification and characterization of a novel G protein coupled receptor, Mas-Related G-Protein-coupled Receptor X2 (MRGPRX2). This receptor promotes pseudoallergic reaction to U.S. Food and Drug Administration (FDA) approved drugs and chronic inflammation in diseases such as asthma, rosacea and hives (urticaria). While the role of MRGPRX2 in promoting allergic reactions has been investigated, the molecular mechanisms utilized by this receptor is poorly understood. Our data suggests that a Ca2+ sensor protein, stromal interaction molecule 1 (STIM1), which is an essential component of the store operated Ca2+ entry (SOCE) pathway, regulates MRGPRX2- and MrgprB2- (the mouse ortholog of the human receptor) induced responses in mast cells. Specifically, silencing the expression of STIM1 or pharmacological targeting of SOCE pathway results in reduced intracellular Ca2+ mobilization, degranulation and cytokine release following mast cell activation via MRGPRX2/MrgprB2 in vitro. Consistent with this data, inflammation is also reduced in in vivo mouse models of rosacea and paw edema when the SOCE pathway is pharmacologically inhibited. This proposal builds up on these observations; the central hypothesis is that STIM1 is a critical regulator of MRGPRX2 responses in mast cells. We will determine the role of STIM1 in regulating human MRGPRX2 and mouse MrgprB2 response in vitro and in vivo (Aim 1). STIM1 couples to the Orai1 channel to mediate SOCE in several cell types. In Aim 1, we will also test whether the STIM1-Orai1 interaction regulates mast cells responses following MRGPRX2/MrgprB2 activation. Similar to data obtained with STIM1 knockdown mast cells, silencing the expression of Na+/H+ exchanger regulatory factor (NHERF)1, an adaptor protein reduced MRGPRX2/MrgprB2-dependent Ca2+ mobilization in vitro and passive systemic anaphylaxis in vivo. How NHERF1 regulates STIM1 functions and affect intracellular Ca2+ mobilization is currently unknown. In Aim 2, we will explore the contribution of the NHERF1-STIM1-Orai1 axis in modulating mast cell response via the MRGPRX2/MrgprB2 receptors. Given the critical role of mast cell MRGPRX2 in causing pseudoallergic reactions and chronic inflammation in allergic diseases, elucidation of the mechanisms by which STIM1 and NHERF1 regulates mast cell MRGPRX2-mediated allergic response is of significant scientific and clinical importance. The successful completion of the proposed studies will likely lead to the identification of potential targets of the MRGPRX2 pathway that can provide insights into the future development of drugs for not only pseudoallergic reactions but also other mast cell-mediated inflammatory diseases.

项目摘要 肥大细胞是组织驻留的先天性免疫细胞，其最佳特征在于其在介导免疫应答中的作用。 过敏性疾病肥大细胞研究的最新进展是鉴定和表征肥大细胞， 一种新的G蛋白偶联受体，Mass相关G蛋白偶联受体X2（MRGPRX 2）。该受体 促进对美国食品和药物管理局（FDA）批准的药物的假性过敏反应， 炎症的疾病，如哮喘，红斑痤疮和荨麻疹（荨麻疹）。虽然MRGPRX 2在促进 尽管已经研究了过敏反应，但是对该受体所利用的分子机制知之甚少。 我们的数据表明，钙离子传感器蛋白，基质相互作用分子1（STIM 1），这是一个必不可少的 钙库操纵的Ca 2+进入（SOCE）途径的组分，调节MRGPRX 2-和MrgprB 2-（小鼠 人受体的直系同源物）在肥大细胞中诱导应答。具体地说，沉默STIM 1的表达 或SOCE途径的药理学靶向导致细胞内Ca 2+动员减少，脱粒 和体外肥大细胞通过MRGPRX 2/MrgprB 2活化后的细胞因子释放。与此数据一致， 当SOCE途径被阻断时， 很压抑。这项提议建立在这些观察的基础上;中心假设是STIM 1 是肥大细胞中MRGPRX 2反应的关键调节因子。我们将确定STIM 1在调节 人MRGPRX 2和小鼠MrgprB 2体外和体内应答（目的1）。STIM 1与Orai 1耦合 通道介导几种细胞类型的SOCE。在目标1中，我们还将测试STIM 1-Orai 1相互作用是否 调节MRGPRX 2/MrgprB 2激活后的肥大细胞反应。与STIM 1获得的数据相似 敲低肥大细胞，沉默Na+/H+交换调节因子（NHERF）1的表达， 蛋白减少体外MRGPRX 2/MrgprB 2依赖性Ca 2+动员和 vivo. NHERF 1如何调节STIM 1功能并影响细胞内Ca 2+动员目前尚不清楚。在 目的2，我们将探索NHERF 1-STIM 1-Orai 1轴在调节肥大细胞反应中的作用， MRGPRX 2/MrgprB 2受体。鉴于肥大细胞MRGPRX 2在引起假性过敏中的关键作用， 反应和慢性炎症的过敏性疾病，阐明的机制，STIM 1和 NHERF 1调节肥大细胞MRGPRX 2介导的过敏反应具有重要的科学和临床意义 重要性成功完成拟议的研究将可能导致确定潜在的 MRGPRX 2通路的靶点，不仅可以为药物的未来发展提供见解， 假性过敏反应，以及其他肥大细胞介导的炎性疾病。