H1R Signaling and Immune Deviation in EAE

EAE 中的 H1R 信号传导和免疫偏差

• 批准号: 8015313
• 负责人: CORY TEUSCHER
• 金额: $ 32.7万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-19 至 2014-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8015313
• 关键词: Adjuvant; Affect; Alleles; Amines; Amino Acids; Arrestins; Autoimmune Diseases; Autoimmune Process; Bordetella pertussis; CD4 Positive T Lymphocytes; Candidate Disease Gene; Cell surface; Cells; Central Nervous System Diseases; Chimeric Proteins; Complement; Data; Development; Disease; Disease Resistance; Disease susceptibility; Endoplasmic Reticulum; Enhancers; Exhibits; Experimental Autoimmune Encephalomyelitis; Exposure to; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Gene Cluster; Gene Transfer Techniques; Genes; Genetic; Genetic Polymorphism; Histamine; Histamine H1 Receptors; Hypersensitivity; Immune; Immune response; In Vitro; Inbred Strains Mice; Inflammation Mediators; Inflammatory; Interferons; Interleukin-17; Knowledge; Lead; Ligand Binding; Link; Linkage Disequilibrium; Maps; Mediating; Mitogen-Activated Protein Kinases; Modeling; Molecular Chaperones; Multiple Sclerosis; Mus; Mutation; Pathway interactions; Peptides; Pertussis Toxin; Phenotype; Play; Predisposition; Process; Production; Proteins; Receptor Activation; Receptor Signaling; Regulation; Resistance; Role; SJL/J Mouse; Signal Pathway; Signal Transduction; Signaling Protein; Surface; T-Lymphocyte; Transfection; Ursidae Family; central nervous system demyelinating disorder; cytokine; gene cloning; human MAPK14 protein; immune function; in vivo; loss of function; mouse model; mutant; novel; novel therapeutics; oligodendrocyte-myelin glycoprotein; overexpression; protein activation; protein folding; protein transport; public health relevance; receptor expression; research study; restoration; trafficking

DESCRIPTION (provided by applicant): Histamine is a potent mediator of inflammation and regulator of innate and adaptive immune responses, and autoimmune diseases such as experimental allergic encephalomyelitis (EAE), the principal autoimmune model of multiple sclerosis (MS). EAE is an inflammatory demyelinating disease of the central nervous system with a critical genetic component. We identified Bphs as a susceptibility locus for EAE and positional candidate gene cloning demonstrated Bphs to be the histamine H1 receptor (Hrh1/H1R). As in many autoimmune diseases, CD4+ T cells play a key role in MS and EAE through production of cytokines such as interferon-g and interleukin-17. T cell-specific expression of the susceptible (BphsS/H1RS) allele allowed for EAE development on a H1RKO background, while expression of the resistant (BphsR/H1RR) allele in the same fashion did not. H1R alleles differ by three amino acids in the third intracellular loop (H1RS: Pro263Val312Pro330; H1RR: Leu263Met312Ser330). This domain is associated with signal transduction, but may also be important in protein folding and trafficking, like the intracellular domains of other G-protein coupled receptors (GPCRs). In transfection studies, H1RR surface expression was substantially lower than H1RS, with H1RR exhibiting intracellular retention in the endoplasmic reticulum (ER). In Aim 1 we will dissect the mechanism(s) responsible for the ER-retention of the H1RR allele and whether pharmacological chaperones can restore H1RR surface expression and affect disease in vivo. Pharmacological chaperones restore surface expression and function of polymorphic GPCRs in vitro, and knowledge gained from the proposed in vivo experiments may have broad implications for GPCR-diseases. We also have evidence of a functional, complementary mechanism that can overcome the ER retention process in BphsR mice. Compilation of Hrh1 sequence data and Bphs data from ~100 inbred mouse strains, revealed strains that were phenotypically BphsS, but bear the H1RR allele. Preliminary mapping studies linked the locus to Hrh1 and because it corrects BphsR, we refer to this gene as Bphs-enhancer (Bphse). SJL/J mice exhibit another Hrh1-linked phenotype, spontaneous histamine sensitivity (Shs). We hypothesize that Bphse and Shs reside within a functional linkage disequilibrium (LD)-domain linked to Hrh1 and may contain genes related to H1R folding, trafficking, or signaling. In Aim 2 we will use a forward genetics approach to positionally clone and identify Bphse and Shs. H1R signaling through p38 mitogen activated protein kinase (MAPK) is important in EAE susceptibility. For many GPCRs sustained MAPK activation involves non-G-protein signaling through b-arrestins. Since we found no difference in G-protein activation of allelic H1R-Ga fusion proteins, this suggested that H1R signaling may also involve a novel non-G-protein b-arrestin pathway. Thus in Aim 3 we will examine whether non-G-protein signaling downstream of the H1R is required for EAE development. Non-G-protein signaling by GPCRs is an emerging paradigm and new pharmacologics are being developed to selectively target these pathways. PUBLIC HEALTH RELEVANCE: The function of G-protein coupled receptors (GPCRs) such as the histamine H1 receptor (H1R) can be regulated by their subcellular localization and the signaling pathways they elicit. Proper folding and cell surface expression of GPCRs is required for ligand binding and signaling. Mutations that lead to improper folding and/or intracellular trafficking comprise the largest class of GPCR mutations that result in disease. Our finding that H1R alleles controlling susceptibility to autoimmune disease exhibit differential cell surface expression and altered intracellular trafficking, with the resistant allele being retained within the endoplasmic reticulum, was the first demonstration that polymorphisms influencing GPCR trafficking and cell surface expression can regulate immune functions. Understanding the mechanisms leading to differences in the trafficking and cell surface expression of the H1R alleles, and how they can be manipulated in vivo using this naturally occurring mouse model, will undoubtedly aid in the development of new therapeutic strategies for diseases in which gain- or loss-of-function mutants leading to GPCR misfolding and/or improper intracellular trafficking are implicated.

描述（由申请人提供）：组胺是一种有效的炎症介质，是先天性和适应性免疫应答以及自身免疫性疾病（如实验性过敏性脑脊髓炎（EAE），多发性硬化症（MS）的主要自身免疫模型）的调节剂。EAE是一种中枢神经系统的炎性脱髓鞘疾病，具有关键的遗传成分。我们确定Bphs作为EAE的易感位点和位置候选基因克隆证明Bphs是组胺H1受体（Hrh 1/H1 R）。与许多自身免疫性疾病一样，CD 4 + T细胞通过产生细胞因子如干扰素-g和白细胞介素-17在MS和EAE中发挥关键作用。易感（BphsS/H1 RS）等位基因的T细胞特异性表达允许在H1 RKO背景下EAE的发展，而抗性（BphsR/H1 RR）等位基因以相同的方式表达则不允许。H1 R等位基因在第三胞内环中相差三个氨基酸（H1 RS：Pro263 Val 312 Pro330; H1 RR：Leu 263 Met 312 Ser 330）。该结构域与信号转导相关，但也可能在蛋白质折叠和运输中很重要，如其他G蛋白偶联受体（GPCR）的胞内结构域。在转染研究中，H1 RR表面表达显著低于H1 RS，H1 RR表现出内质网（ER）中的细胞内滞留。在目标1中，我们将剖析负责H1 RR等位基因的ER保留的机制，以及药理学伴侣是否可以恢复H1 RR表面表达并影响体内疾病。药理学伴侣在体外恢复多态性GPCR的表面表达和功能，并且从所提出的体内实验中获得的知识可能对GPCR疾病具有广泛的影响。我们也有一个功能性的，互补的机制，可以克服在BphsR小鼠的ER保留过程的证据。来自约100个近交系小鼠品系的Hrh 1序列数据和Bphs数据的汇编揭示了表型为BphsS但携带H1 RR等位基因的品系。初步的定位研究将该基因座与Hrh 1联系起来，因为它纠正了BphsR，我们将该基因称为Bphs-enhancer（Bphse）。SJL/J小鼠表现出另一种Hrh 1连锁表型，自发组胺敏感性（Shs）。我们假设Bphse和Shs位于与Hrh 1相关的功能性连锁不平衡（LD）结构域内，可能包含与H1 R折叠、运输或信号传导相关的基因。在目标2中，我们将使用正向遗传学方法定位克隆和鉴定Bphse和Shs。通过p38丝裂原活化蛋白激酶（MAPK）的H1 R信号传导在EAE易感性中是重要的。对于许多GPCR，持续的MAPK激活涉及通过b-抑制蛋白的非G蛋白信号传导。由于我们发现等位基因H1 R-Ga融合蛋白的G蛋白活化没有差异，这表明H1 R信号传导也可能涉及新的非G蛋白b-抑制蛋白途径。因此，在目标3中，我们将研究H1 R下游的非G蛋白信号传导是否是EAE发展所必需的。通过GPCR进行的非G蛋白信号传导是一种新兴的范式，并且正在开发新的药理学来选择性地靶向这些途径。 公共卫生关系：G蛋白偶联受体（GPCR）如组胺H1受体（H1 R）的功能可通过其亚细胞定位和它们引发的信号传导途径来调节。GPCR的正确折叠和细胞表面表达是配体结合和信号传导所需的。导致不正确折叠和/或细胞内运输的突变包括导致疾病的最大类别的GPCR突变。我们的发现，H1 R等位基因控制自身免疫性疾病的易感性表现出不同的细胞表面表达和改变细胞内的运输，与抗性等位基因被保留在内质网内，是第一个证明多态性影响GPCR运输和细胞表面表达可以调节免疫功能。了解机制导致的差异，在运输和细胞表面表达的H1 R等位基因，以及它们如何可以在体内使用这种天然存在的小鼠模型进行操纵，无疑将有助于开发新的治疗策略的疾病，其中获得或丧失功能的突变体，导致GPCR错误折叠和/或不适当的细胞内运输牵连。