Regulation of Toll-like receptor-induced NFkB activation by Gai-coupled receptors

描述（由申请人提供）：我的职业目标是指导一个独立的研究实验室，探索介导宿主防御和炎症的分子机制。在炎症反应期间，G蛋白偶联受体（GPCR）和先天免疫受体同时被激活，并且各自可以激活不同的细胞信号。总体假设是炎性GPCR信号传导与巨噬细胞Toll样受体（TLR）活化途径的许多组分相互作用并修饰巨噬细胞Toll样受体（TLR）活化途径的许多组分，最终增强促炎性转录因子核因子κ B（NF κ B）的活化。这得到了初步发现的支持，即白三烯B4（LTB 4）连接其GPCR BLT 1增强了NFkB的TLR依赖性活化。GPCR和TLR信号传导之间存在许多潜在的相互作用，其可能组合地决定NF κ B活化的状态。为了发现这样的相互作用，我们提出了一个跨学科的方法结合实验与系统生物学模型的综合网络，能够被实验tested.The总体计划的建议是获得知识和技能，在我最初两年的指导培训使用BLT 1-TLR相互作用作为一个模型，这将是在独立阶段扩展到其他类的GPCR。为了验证我们的假设，提出了以下具体目标：K99期;目标1：确定LTB 4/BLT 1信号传导的重要性（激酶，ROI生成）对TLR依赖性NF κ B活化的影响，目的2：检查LTB 4/BLT 1信号传导对TLR/IL-1 β受体及其衔接子表达的影响，目的3：开发计算模型（方程），涉及在Mo TLR反应期间由BLT 1诱导的信号程序。R 00期：目标4：基于在K99期间获得的发现，我们想要进一步研究除了BLT 1之外的Gai蛋白偶联受体在TLR诱导的NFkB活化中的重要性。目的5：计算模型调节NFkB活化的细胞信号传导网络（GPCR和TLR），以便制定可检验的假设。这个提议将为炎症中巨噬细胞激活的协调提供新的见解，并将促进我成为一名独立的研究者。 公共卫生相关性：我们的提议具有直接的翻译重要性，因为GPCR、TLR和NF κ B是治疗干预的主要靶点，并且数学模型的开发可以通过采用抑制剂阻断炎症反应以及通过添加外源性GPCR激动剂增强炎症来解锁治疗压倒性以及免疫抑制状态的更具体的策略。

DESCRIPTION (provided by applicant): My career goal is to direct an independent research laboratory exploring the molecular mechanisms mediating host defense and inflammation. During inflammatory responses, both G protein-coupled (GPCRs) and innate immune receptors are activated simultaneously, and each can activate diverse cellular signals. The overall hypothesis is that inflammatory GPCR signaling interacts with and modifies many components of the macrophage Toll-like receptor (TLR) activation pathway, ultimately enhancing activation of the pro-inflammatory transcription factor nuclear factor kappa B (NFkB). This is supported by the preliminary finding that leukotriene B4 (LTB4) ligation of its GPCR BLT1 enhances the TLR- dependent activation of NFkB. There are many potential interactions between GPCR and TLR signaling that might combinatorially determine the state of NFkB activation. To discover such interactions, we propose an interdisciplinary approach combining experimentation with systems biology to model integrative networks that are capable of being tested experimentally.The general plan of this proposal is to acquire knowledge and skills during my initial two years of mentored training using BLT1-TLR interactions as a model, which will be extended to other classes of GPCRs during the independent phase. To test our hypothesis, the following Specific Aims are proposed: K99 period; Aim 1: Determine the importance of LTB4/BLT1 signaling (kinases, ROI generation) to TLR-dependent NFkB activation, Aim 2: Examine the effects of LTB4/BLT1 signaling on the expression of TLRs/IL-1beta receptors and their adaptors, Aim 3: Develop computational models (equations) involving the signaling programs induced by BLT1 during Mo TLR responses. R00 period:Aim 4: Based on the findings obtained in the K99 period, we want to further investigate the importance of Gai protein-coupled receptors other than BLT1 in TLR-induced NFkB activation, Aim 5: Computationally model cellular signaling networks (GPCR and TLRs) that regulate NFkB activation in order to formulate testable hypotheses. This proposal will provide new insights into the coordination of macrophage activation in inflammation, and will foster my development into an independent investigator. PUBLIC HEALTH RELEVANCE: Our proposal have direct translational importance, since GPCRs, TLRs and NFkB are major targets of therapeutic interventions and the development of mathematical models could unlock more specific strategies to treat both overwhelming as well as immunosuppressive states by employing inhibitors to block inflammatory responses, as well as by adding exogenous GPCR agonists to enhance inflammation.