The Structure and Function of the CRTH2 PDG2 Receptor

描述（由申请人提供）： 前列腺素D2（PGD 2）是一种主要的环氧合酶代谢产物，其释放被认为是哮喘中观察到的炎症和气道高反应性增加的原因。PGD 2似乎部分通过最近克隆的GPCR（命名为CRTH 2）发挥作用。CRTH 2 mRNA表达模式与该受体在过敏性疾病病因学中的重要作用一致。 该提议的主要假设是，免疫炎症反应中关键的PGD 2诱发的变化至少部分地由CRTH 2诱发的反应介导。我们进一步提出，PGD 2代谢产物（包括PGJ 2和15 d PGJ 2）的一些作用是通过CRTH 2受体介导的。 为了检验这一假设，我们在具体目标1中提出定义小鼠CRTH 2受体的药理学。我们将在细胞培养中表达CRTH 2受体，并确定其配体结合和信号转导特性。在特定目标2中，我们将通过定点诱变鉴定对配体结合和信号转导至关重要的氨基酸残基。使用建模和分子动力学模拟作为指导，我们将引入点突变CRTH 2受体，并在细胞培养系统中表达突变受体。在具体目标3中，我们将探讨PGD 2是否通过CRTH 2受体调节哮喘小鼠模型的炎症效应和/或支气管收缩。我们将在CRTH 2基因敲除小鼠中分析卵清蛋白（OVA）过敏性致敏（TH 2型免疫模型），并将其与DP无效动物进行比较。我们进一步提出将CRTH 2缺失小鼠与DP缺失小鼠杂交，以测试两种PGD 2受体缺失的影响。这些研究的完成应有助于阐明PGD 2在过敏性气道疾病中细胞迁移和浸润中的生理作用，并将提供关于该受体在小鼠过敏性炎症模型中的作用的见解。

DESCRIPTION (provided by applicant): Prostaglandin D2 (PGD2) is a major cyclooxygenase metabolite, and its release has been hypothesized to contribute to the inflammation and increased airway hyperreactivity observed in asthma. PGD2 appears to act in part through a recently cloned GPCR designated CRTH2. CRTH2 mRNA expression pattern is consistent with an important role for this receptor in the etiology of allergic disease. The principal hypothesis of this proposal is that critical PGD2 evoked changes in the immune inflammatory response are mediated, at least in part, by CRTH2-evoked responses. We further propose that some of the effects of PGD2 metabolites including PGJ2 and 15d PGJ2 are mediated via the CRTH2 receptor. To test this hypothesis we propose in Specific Aim 1 to define the pharmacology of the mouse CRTH2 receptor. We will express the CRTH2 receptor in cell culture and determine its ligand binding and signal transduction properties. In Specific Aim 2 we will identify amino acid residues critical for ligand binding and signal transduction by site directed mutagenesis. Using modeling and molecular dynamics simulations as a guide, we will introduce point mutations into the CRTH2 receptor, and express mutant receptors in cell culture systems. In Specific Aim 3 we will explore whether PGD2 modulates inflammatory effects and/or bronchoconstriction in mouse models of asthma via the CRTH2 receptor. We will analyze ovalbumin (OVA) allergic sensitization, a TH2 type immune model, in CRTH2 knockout mice and compare this with DP null animals. We further propose to cross the CRTH2 null mice with DP null mice to test the effect of loss of both PGD2 receptors. Completion of these studies should aid in the elucidation of the physiologic role of PGD2 in cellular migration and infiltration in allergic airway disease and will provide insight as to the role of this receptor in mouse models of allergic inflammation.