Signaling to PIAS1 to Regulate Immune Responses

描述（由申请人提供）：炎症刺激引发的免疫反应必须严格调节。无限制的炎症与免疫紊乱有关。细胞外炎症刺激如何向细胞核发出信号以限制炎症基因激活尚不清楚。NF吗?B和STATs是两个重要的转录因子家族，它们被各种促炎刺激激活以诱导基因表达。活化STAT1蛋白抑制剂（PIAS1）通过选择性阻断NF？B和STAT1到基因启动子。我们最近发现了一种新的信号通路，在这种通路中，促炎刺激激活ikka介导的PIAS1磷酸化，从而立即抑制炎症基因的激活。本研究计划的总体目标是表征IKKa-PIAS1信号通路在免疫应答调节中的作用。具体来说，我们将通过突变分析和激酶测定来表征ikka介导的PIAS1 Ser90磷酸化的分子基础。IKK家族其他成员的作用以及IKK在介导PIAS1 Ser90磷酸化中的特异性将通过IKK敲除细胞进行检测。接下来，我们将研究PIAS1磷酸化如何调节PIAS1的活性。我们将研究PIAS1 Ser90磷酸化在配体诱导的PIAS1启动子募集中的作用。我们将分析PIAS1 SAP结构域在Ser90磷酸化依赖性PIAS1启动子募集中的作用。我们将测试Ser90磷酸化是否足以使PIAS1靶向基因启动子，使用Ser90磷酸化突变体。PIAS1的启动子结合区(s)将通过ChIP检测和突变分析进行检测。最后，我们将构建PIAS1磷酸化缺陷敲入小鼠模型，研究PIAS1磷酸化在病原感染应答中的生理功能。这些研究将探索免疫反应负调控的一种新的分子信号机制，这将提高我们设计合理治疗感染性和炎症性疾病的能力。与公共卫生相关：必须严格调节炎症刺激引发的免疫反应。无限制的炎症与免疫系统紊乱和癌症有关。本研究旨在研究一种新发现的平衡免疫应答的信号通路。这些研究将增强我们设计治疗免疫疾病的新治疗策略的能力。

DESCRIPTION (provided by applicant): The immune responses triggered by inflammatory stimuli must be tightly regulated. Unrestricted inflammation is associated with immune disorders. How extracellular inflammatory stimuli signal to the nucleus to restrict inflammatory gene activation is poorly understood. NF?B and STATs are two important families of transcription factors that are activated by a wide variety of pro-inflammatory stimuli to induce gene expression. Protein inhibitor of activated STAT1 (PIAS1) inhibits immune responses by selectively blocking the binding of NF?B and STAT1 to gene promoters. We have recently identified a novel signaling pathway in which pro- inflammatory stimuli activate the IKKa-mediated phosphorylation of PIAS1 for the immediate repression of inflammatory gene activation. The overall goal of this research proposal is to characterize the IKKa-PIAS1 signaling pathway in the regulation of immune responses. Specifically, we will characterize the molecular basis of the IKKa-mediated PIAS1 Ser90 phosphorylation by mutational analysis and kinase assays. The role of other members of the IKK family and the specificity of IKKs in mediating PIAS1 Ser90 phosphorylation will be examined using IKK knockout cells. Next, we will investigate how PIAS1 phosphorylation regulates the activity of PIAS1. We will examine the role of PIAS1 Ser90 phosphorylation in the ligand-induced promoter recruitment of PIAS1. We will analyze the role of PIAS1 SAP domain in the Ser90 phosphorylation-dependent promoter recruitment of PIAS1. We will test if Ser90 phosphorylation is sufficient to target PIAS1 to gene promoters using Ser90 phosphomimic mutants. The promoter binding region(s) of PIAS1 will be examined by ChIP assays and mutational analysis. Finally, we will generate a PIAS1 phosphorylation-defective knockin mouse model to study the physiological function of PIAS1 phosphorylation in response to pathogenic infection. These studies will investigate a novel molecular signaling mechanism for the negative regulation of immune responses, which will enhance our ability to design rational therapeutic strategies for the treatment of infectious and inflammatory diseases. RELEVANCE TO PUBLIC HEALTH: The immune responses triggered by inflammatory stimuli must be tightly regulated. Unrestricted inflammation is associated with immune disorders and cancer. This proposal is to study a newly identified signaling pathway that functions to balance immune responses. These studies will enhance our ability to design novel therapeutic strategies for the treatment of immune diseases.