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.

项目概要 肥大细胞是组织驻留的先天免疫细胞，其在介导中的作用得到了最好的表征。 过敏性疾病。肥大细胞研究的最新进展是肥大细胞的鉴定和表征 一种新型 G 蛋白偶联受体，Mas 相关 G 蛋白偶联受体 X2 (MRGPRX2)。该受体 促进对美国食品和药物管理局 (FDA) 批准的药物和慢性药物的假过敏反应 哮喘、红斑痤疮和荨麻疹（荨麻疹）等疾病引起的炎症。而 MRGPRX2 在促进 过敏反应已被研究，但对该受体利用的分子机制知之甚少。 我们的数据表明 Ca2+ 传感器蛋白，基质相互作用分子 1 (STIM1)，它是一种重要的 钙池操作 Ca2+ 进入 (SOCE) 途径的组成部分，调节 MRGPRX2- 和 MrgprB2-（小鼠 人类受体的直系同源物）诱导肥大细胞的反应。具体来说，沉默 STIM1 的表达 SOCE 途径的药理学靶向导致细胞内 Ca2+ 动员、脱颗粒减少 体外通过 MRGPRX2/MrgprB2 激活肥大细胞后细胞因子的释放。与此数据一致， 当 SOCE 途径被激活时，体内红斑痤疮和爪水肿小鼠模型中的炎症也会减少 药理学上有抑制作用。该提案建立在这些观察的基础上；中心假设是 STIM1 是肥大细胞中 MRGPRX2 反应的关键调节因子。我们将确定 STIM1 在调节中的作用 人 MRGPRX2 和小鼠 MrgprB2 的体外和体内反应（目标 1）。 STIM1 与 Orai1 耦合 通道介导多种细胞类型中的 SOCE。在目标 1 中，我们还将测试 STIM1-Orai1 是否相互作用 调节 MRGPRX2/MrgprB2 激活后肥大细胞的反应。与 STIM1 获得的数据类似 敲低肥大细胞，沉默 Na+/H+ 交换调节因子 (NHERF)1（一种接头）的表达 蛋白质减少了 MRGPRX2/MrgprB2 依赖性 Ca2+ 体外动员和被动全身过敏反应 体内。 NHERF1 如何调节 STIM1 功能并影响细胞内 Ca2+ 动员目前尚不清楚。在 目标 2，我们将通过以下方式探索 NHERF1-STIM1-Orai1 轴在调节肥大细胞反应中的贡献 MRGPRX2/MrgprB2 受体。鉴于肥大细胞 MRGPRX2 在引起假性过敏中的关键作用 过敏性疾病中的反应和慢性炎症，阐明 STIM1 和 NHERF1调节肥大细胞MRGPRX2介导的过敏反应具有重要的科学和临床意义 重要性。拟议研究的成功完成可能会导致潜在的识别 MRGPRX2 通路的靶标不仅可以为药物的未来开发提供见解 假性过敏反应以及其他肥大细胞介导的炎症性疾病。