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

描述（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。在炎症反应中，G蛋白偶联（gpcr）和先天免疫受体同时被激活，并且各自可以激活多种细胞信号。总体假设是炎症性GPCR信号与巨噬细胞toll样受体（TLR）激活途径的许多组分相互作用并修饰，最终增强促炎转录因子核因子κ B （NFkB）的激活。初步发现白三烯B4 （LTB4）连接其GPCR BLT1可增强NFkB的TLR依赖性激活。GPCR和TLR信号之间存在许多潜在的相互作用，这些相互作用可能共同决定NFkB的激活状态。为了发现这种相互作用，我们提出了一种跨学科的方法，将实验与系统生物学相结合，以模拟能够通过实验测试的综合网络。本次提案的总体计划是在前两年的指导培训中，以BLT1-TLR相互作用为模型，学习知识和技能，并在独立阶段扩展到其他类别的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LTB4/BLT1信号（激酶，ROI生成）对TLR依赖性NFkB激活的重要性；目标2：检查LTB4/BLT1信号对TLRs/ il -1 β受体及其接头表达的影响；目标3：建立涉及BLT1在Mo TLR反应中诱导的信号程序的计算模型（方程）。目标4：基于K99期的发现，我们希望进一步研究除BLT1外的Gai蛋白偶联受体在tlr诱导的NFkB激活中的重要性。目标5：计算模拟调节NFkB激活的细胞信号网络（GPCR和TLRs），以制定可测试的假设。这一提议将为炎症中巨噬细胞活化的协调提供新的见解，并将促进我成为一名独立的研究者。

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.