NHERF1 regulates MRGPRX2/MrgprB2 responses in mast cells

NHERF1 调节肥大细胞中的 MRGPRX2/MrgprB2 反应

• 批准号: 10711042
• 负责人: Hariharan Subramanian
• 金额: $ 46.07万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-12 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10711042
• 关键词: Adaptor Signaling Protein; Affect; Agonist; Allergic; Allergic Disease; Allergic Reaction; Anaphylaxis; Asthma; Attenuated; Biological Assay; CD34 gene; CISH gene; Cell Compartmentation; Cell physiology; Cells; Cellular biology; Chronic; Clinical; Data; Development; Disease; Edema; Emergency department visit; Engraftment; Expenditure; Female; Future; G-Protein-Coupled Receptors; Goals; Granulocyte-Macrophage Colony-Stimulating Factor; Healthcare; Human; IL3 Gene; IgE Receptors; Immune; In Vitro; Inflammation; Inflammatory; Intervention; Ligands; Mediating; MicroRNAs; Molecular; Mouse Strains; Mus; Orthologous Gene; Pathogenesis; Pathway interactions; Pharmaceutical Preparations; Phosphorylation; Play; Proteins; Proteomics; Reaction; Regulation; Research; Role; Rosacea; Signal Pathway; Signal Transduction; Signaling Molecule; Signaling Protein; Site; Testing; Time; Tissues; Umbilical Cord Blood; United States Food and Drug Administration; Urticaria; allergic response; cathelicidin; cathelicidin antimicrobial peptide; chronic inflammatory disease; clinical application; cytokine; drug development; experimental study; humanized mouse; in vivo; in vivo Model; inositol-1,4,5-trisphosphate 5-phosphatase; insight; mast cell; mastocytosis; mimetics; mortality; mouse model; mutant; novel; novel therapeutics; overexpression; pharmacologic; phosphoproteomics; receptor; response; sodium-hydrogen exchanger regulatory factor; stem cells; transcriptomics

PROJECT SUMMARY Mast cells are tissue resident innate immune cells that have been best characterized for their role in mediating allergic diseases. A new 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 thoroughly investigated, the molecular mechanisms utilized by this receptor is poorly understood. Our data suggests that Na+/H+ exchanger regulatory factor (NHERF)1, an adaptor protein, regulates MRGPRX2- and MrgprB2- (the mouse ortholog of the human receptor) induced responses in mast cells. Specifically, reducing the expression of NHERF1 in mast cells results in decreased intracellular Ca2+ mobilization and degranulation via MRGPRX2/MrgprB2 in vitro. Consistent with this data, NHERF1 also promoted anaphylaxis in MrgprB2-dependent mouse models of paw edema and passive systemic anaphylaxis in vivo. This proposal builds up on these observations; the central hypothesis is that NHERF1 is a critical regulator of MRGPRX2 responses in mast cells. Using a novel mouse strain that we have recently generated, Cpa3- Cre+:NHERF1fl/fl, primary human CD34+ stem cell-derived mast cells and humanized mice, we will determine the role of mast cell-specific expression of NHERF1 in regulating mouse MrgprB2 and human MRGPRX2 receptors in vitro and in vivo (Aim 1). In Aim 2, we will explore the mechanisms through which NHERF1 modulates mast cell response. Specifically, we will determine whether phosphorylation of NHERF1 is important for NHERF1 regulation of MRGPRX2/MrgprB2. We will identify the components of the NHERF1-interactome and signaling proteins and pathways through which NHERF1 functions in mast cells by using transcriptomic and proteomic approaches. Additionally, we will delineate the role of NHERF1-microRNA (miR)155-suppressor of cytokine signaling (SOCS1)-Src homology 2 (SH2) domain containing inositol polyphosphate 5-phosphatase 1 (SHIP-1) axis in promoting mast cell activation. Given the critical role of mast cell-MRGPRX2 in causing pseudoallergic reactions and chronic inflammation in allergic diseases, elucidation of the mechanisms by which NHERF1 contributes to MRGPRX2-mediated allergic response is of significant scientific and clinical importance. We firmly believe that 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。肥大细胞研究的一个新进展是肥大细胞的鉴定和表征 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 过敏反应已被彻底研究，该受体利用的分子机制还很差 明白了。我们的数据表明 Na+/H+ 交换调节因子 (NHERF)1（一种接头蛋白）可调节 MRGPRX2- 和 MrgprB2-（人类受体的小鼠直系同源物）诱导肥大细胞的反应。 具体而言，减少肥大细胞中 NHERF1 的表达会导致细胞内 Ca2+ 动员减少 并通过 MRGPRX2/MrgprB2 进行体外脱颗粒。与此数据一致，NHERF1 还推广了 MrgprB2 依赖性爪水肿小鼠模型中的过敏反应和体内被动全身过敏反应。这 提案建立在这些观察的基础上；中心假设是 NHERF1 是 肥大细胞中的 MRGPRX2 反应。使用我们最近培育的新型小鼠品系，Cpa3- Cre+:NHERF1fl/fl，原代人类 CD34+ 干细胞衍生的肥大细胞和人源化小鼠，我们将确定 NHERF1 肥大细胞特异性表达在调节小鼠 MrgprB2 和人 MRGPRX2 受体中的作用 体外和体内（目标 1）。在目标 2 中，我们将探索 NHERF1 调节桅杆的机制 细胞反应。具体来说，我们将确定 NHERF1 的磷酸化对于 NHERF1 是否重要 MRGPRX2/MrgprB2 的调节。我们将鉴定 NHERF1 相互作用组和信号传导的组成部分 通过转录组学和蛋白质组学研究 NHERF1 在肥大细胞中发挥作用的蛋白质和途径 接近。此外，我们将描述 NHERF1-microRNA (miR)155-细胞因子抑制剂的作用 含有肌醇多磷酸 5-磷酸酶 1 (SHIP-1) 的信号转导 (SOCS1)-Src 同源 2 (SH2) 结构域 轴促进肥大细胞活化。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 NHERF1 的机制 促进 MRGPRX2 介导的过敏反应具有重要的科学和临床意义。我们坚决 相信成功完成拟议的研究将可能导致潜在目标的确定 MRGPRX2 通路的研究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。