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Role of beta-arrestin-2 on IgE-mediated cofilin dephosphorylation and mast cell activation

beta-arrestin-2 对 IgE 介导的丝切蛋白去磷酸化和肥大细胞激活的作用

基本信息

批准号：
9114460
负责人：
Hydar Ali
金额：
$ 24万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-01 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9114460
关键词：
ADRBK1 gene AIDS/HIV problem ARRB1 gene ARRB2 Actins Adrenal Cortex Hormones Affinity Agonist Allergens Antibodies Antigens Area Arr2 Arrestins Asthma Attenuated Binding Sites Bone Marrow Breeding Carboxypeptidase Cells Chemotaxis Chronic Complement Complement 3a Complex Data Development Disease Extrinsic asthma Family G Protein-Coupled Receptor Signaling G protein coupled receptor kinase G-Protein-Coupled Receptors GTP-Binding Proteins Generations Goals Health Histamine Release Human IgE IgE Receptors Immune response In Vitro Inflammation Mediators Inflammatory Laboratories Leukotriene Antagonists Lipids LoxP-flanked allele Lung Lung diseases MAPK Signaling Pathway Pathway Mediating Medical Morbidity - disease rate Mus Pathogenesis Patients Phosphorylation Phosphotransferases Play Plus End of the Actin Filament Protein Dephosphorylation Protein Kinase Proteins Receptor Signaling Regulation Reporting Retroviridae Role Scaffolding Protein Signal Pathway Signal Transduction Signaling Protein Tertiary Protein Structure Testing Therapeutic Tuberculosis United States antimicrobial peptide base beta-arrestin cofilin cysteinyl-leukotriene cytokine depolymerization desensitization economic cost in vivo lipid mediator mast cell mastocytosis montelukast novel novel strategies novel therapeutic intervention novel therapeutics omalizumab overexpression protein activation receptor response scaffold

项目摘要

DESCRIPTION (provided by applicant): Asthma is one of the most common respiratory diseases world-wide; 300 million people are currently receiving treatment and global economic cost exceeds those of tuberculosis and HIV/AIDS combined. Thus, asthma remains an area of considerable unmet medical need. Allergic asthma is caused by an overzealous Th2 immune response to allergens in which immunoglobulin E (IgE) and mast cells play critical roles. Thus, aggregation of high affinity IgE receptor (FcεRI) by allergen on mast cells results in histamine release and the generation of lipids and cytokines, which are responsible for the manifestations of asthma. The focus of our laboratory has been to study the regulation of G protein coupled receptor (GPCR) signaling in mast cells. Functions of GPCRs are regulated via their phosphorylation by a family of protein kinases known as GPCR kinases (GRKs) and the recruitment of the scaffolding molecule ß-arrestin (ß-arr). The receptor/ß-arr complex serves to attenuate G protein activation leading to receptor desensitization. ß-arr also promotes downstream MAPK signaling pathways independent of G protein activation. To the best of our knowledge, the role of ß-arr on FcεRI signaling in mast cells has never been reported. Based on our unexpected preliminary data, we hypothesize that ß-arr2 serves as a scaffolding protein to promote FcεRI- mediated cofilin dephosphorylation in mast cells. We further hypothesize that this ß-arr2-mediated cofilin dephosphorylation contributes to mast cell recruitment in the lung to promote the development chronic asthma. In aim 1, retrovirus will be used to transduce different domains of ß-arr2 into ß-arr2-/- mast cells and their ability to promote FcεRI-mediated cofilin dephosphorylation and mast cell chemotaxis will be determined. We will then overexpress different sub-domains of ß-arr2 into mast cells endogenously expressing ß-arr2 and their ability to interfere with FcεRI-mediated cofilin dephosphorylation and mast cell chemotaxis will be determined. In aim 2, we will delete ß-arr2 in mast cells by breeding ß-arr2-floxed mice and carboxypeptidase-3 (Cpa-3) cre mice. We will also overexpress ß-arr2 in mast cells in vivo by engrafting ß-arr2-transduced BMMCs into mast cell-deficient Wsh/Wsh mice. These complementary approaches will be used to test the hypothesis that ß-arr2 expressed in mast cells contributes to their recruitment in the lung and promotes the development of chronic asthma. Finally, we will express cofilin and peptide domains of ß-arr2 in mast cells in vivo. These strategies will be used to determine if mast cell-specific expression of cofilin and ß-arr2 domains modulate chronic asthma. This study will likely generate new information regarding the roles of ß-arr2 and cofilin in mast cell chemotaxis in vitro and in vivo and may provide a better rationale for the development of novel therapeutics for chronic asthma.

哮喘是世界范围内最常见的呼吸道疾病之一;目前有3亿人正在接受治疗，全球经济成本超过结核病和艾滋病毒/艾滋病的总和。因此，哮喘仍然是一个相当大的未满足的医疗需求的领域。过敏性哮喘是由对过敏原的过度Th 2免疫应答引起的，其中免疫球蛋白E（IgE）和肥大细胞起关键作用。因此，高亲和力IgE受体（FcεRI）被过敏原聚集在肥大细胞上，导致组胺释放以及脂质和细胞因子的产生，这是导致哮喘表现的原因。本实验室的工作重点是研究肥大细胞中G蛋白偶联受体（GPCR）信号的调控。GPCR的功能通过被称为GPCR激酶（GRKs）的蛋白激酶家族磷酸化和骨架分子β-抑制蛋白（β-arrestin，GRKs）的募集来调节。受体/受体-受体复合物用于减弱G蛋白激活，导致受体脱敏。β-D-半乳糖苷酶还促进下游MAPK信号通路，而不依赖于G蛋白活化。据我们所知，肥大细胞中FcεRI信号通路中的Fc-β受体的作用还未见报道。基于我们意想不到的初步数据，我们假设，在肥大细胞中，α-arr 2作为支架蛋白促进FcεRI介导的cofilin去磷酸化。我们进一步假设，这种p-arr 2介导的cofilin去磷酸化有助于肺中肥大细胞的募集，促进慢性哮喘的发展。在目标1中，将使用逆转录病毒将β-arr 2的不同结构域转染到β-arr 2-/-肥大细胞中，并确定它们促进Fcε RI介导的cofilin去磷酸化和肥大细胞趋化性的能力。然后，我们将在内源性表达α-arr 2的肥大细胞中过表达α-arr 2的不同亚结构域，并测定它们干扰Fcε RI介导的cofilin去磷酸化和肥大细胞趋化性的能力。目的二：通过饲养c-arr 2-floxed小鼠和羧肽酶-3（Cpa-3）cre小鼠，使肥大细胞中的c-arr 2基因缺失。我们还将通过将转导了p53-arr 2的BMMC移植到肥大细胞缺陷型Wsh/Wsh小鼠体内，在肥大细胞中过表达p53-arr 2。这些互补的方法将被用来测试的假设，即肥大细胞中表达的arr 2有助于他们在肺中的招聘，并促进慢性哮喘的发展。最后，我们将在体内肥大细胞中表达cofilin和peptide domains of peptide-arr 2。这些策略将被用来确定是否肥大细胞特异性表达的cofilin和cofil-arr 2结构域调节慢性哮喘。这项研究可能会产生新的信息，在体外和体内肥大细胞趋化性中的作用，arr 2和cofilin，并可能提供一个更好的理由，为慢性哮喘的新疗法的发展。