Rip Proteins in Innate Immune Signaling

项目摘要/摘要 自身免疫性和炎症性疾病发病率的增加突显了改进的必要性 治疗这些疾病的治疗方法。我们的工作将RIPK1激酶置于炎症通路的顶端 它调节一种由RIPK1、RIPK3和RIPK3介导的称为坏死性下垂的炎性细胞死亡 MLKL。在提出的工作中，我们将描绘新的坏死性下垂途径在自身免疫和 自身免疫性和病毒性肝炎的免疫介导性肝损伤手术治疗。 我们证明，具有树突状细胞RIPK1缺陷(Ripk1DC KO)的小鼠会发生炎症和 自身免疫；意外发现RIPK1抑制树突状细胞(DC)的坏死性下垂 它很好地确立了作为坏死性下垂的启动者的作用。在发育过程中，RIPK1可预防坏死性下垂 通过核酸传感器Z-DNA结合蛋白(ZBP1)的介导，我们推测ZBP1- 介导性树突状细胞坏死性下垂破坏耐受性并诱导自身免疫。我们将检验这一假设 目标1通过确定ZBP1缺陷是否能预防Ripk1DC KO的炎症和自身免疫 老鼠。我们将鉴定与ZBP1结合的核酸物种，并将确定核酸结合 是诱导炎症和/或自身免疫所必需的。众所周知，Z形的核酸具有高度的 狼疮患者体内存在免疫原性和抗Z-DNA自身抗体；因此，我们将对其进行基因检测 抑制ZBP1或坏死性下垂是否能改善其他有自身免疫倾向的小鼠的疾病。在目标2中，我们 将研究RIPK1依赖的坏死性下垂是否对自身免疫的小鼠模型有贡献 植物凝集素刀豆球蛋白A(ConA)诱导的肝炎(AIH)。根据我们在年的初步数据 相互的造血嵌合体我们假设AIH是由典型的坏死性下垂介导的 造血细胞与肝细胞非典型性坏死性下垂。与增加的RIPK1和 MLKL在AIH患者肝脏中的表达，我们的初步数据表明RIPK1和MLKL有关，但不是 RIPK3在肝细胞死亡中的作用，使我们假设一种新的RIPK3非依赖性坏死性下垂 途径是在肝细胞中诱导的。下一步，我们将通过生产老鼠来严格检验这一假设 在肝实质细胞中特异表达RIPK1D138N或缺失RIPK3或MLKL的蛋白。一个 另一个目标将是通过鉴定RIPK1-1来描绘这一非典型的坏死性下垂途径。 相互作用激酶负责肝细胞中MLKL的激活，并评估其在AIH中的作用。 这些目标的成功完成将阐明这些新的坏死性下垂途径的贡献(S) 自身免疫和急性肝炎的长期目标是识别这些 可能受益于RIPK1激酶靶向治疗的自体炎症/免疫性疾病

PROJECT SUMMARY/ABSTRACT The increased incidence of autoimmune and inflammatory disorders highlights the need for improved therapeutics to treat these diseases. Our work places RIPK1 kinase at the apex of inflammatory pathways where it regulates a form of inflammatory cell death called necroptosis mediated by RIPK1, RIPK3 and MLKL. In the work proposed, we will delineate novel necroptosis pathways important in autoimmunity and immune-mediated liver injury operative in autoimmune and viral hepatitis. We demonstrate that mice with a dendritic cell RIPK1-deficiency (Ripk1DC KO) develop inflammation and autoimmunity; unexpectedly revealing that RIPK1 restrains necroptosis in dendritic cells (DC) in contrast to it well established role as an initiator of necroptosis. During development, RIPK1 prevents necroptosis mediated by the nucleic acid sensor Z-DNA binding protein (ZBP1) leading us to hypothesize that ZBP1- mediated dendritic cell necroptosis breaks tolerance and induces autoimmunity. We will test this hypothesis in Aim 1 by determining whether a ZBP1-deficiency prevents inflammation and autoimmunity in Ripk1DC KO mice. We will identify the nucleic acid species bound to ZBP1 and will establish whether nucleic acid binding is required to induce inflammation and/or autoimmunity. Nucleic acids in the Z-form are known to be highly immunogenic and anti-Z-DNA autoantibodies occur in lupus patients; therefore, we will test genetically whether inhibiting ZBP1 or necroptosis ameliorates disease in other autoimmune prone mice. In Aim 2, we will investigate whether RIPK1-kinase-dependent necroptosis contributes to a mouse model of autoimmune hepatitis (AIH) induced by the plant lectin Concanavalin A (Con A). Based on our preliminary data in reciprocal hematopoietic chimeras we hypothesize that AIH is mediated by canonical necroptosis in hematopoietic cells and noncanonical necroptosis in hepatocytes. Consistent with increased RIPK1 and MLKL expression in the livers of AIH patients, our preliminary data implicate RIPK1 and MLKL but not RIPK3 in hepatocyte cell death, leading us to hypothesize that a novel RIPK3 independent necroptosis pathway is induced in hepatocytes. Going forward we will test this hypothesis rigorously by generating mice that express kinase inactive RIPK1D138N or delete Ripk3 or Mlkl specifically in liver parenchymal cells. An additional goal will be to delineate this non-canonical necroptosis pathway, by identifying the RIPK1- interacting kinase responsible for MLKL activation in hepatocytes and to assess its contribution to AIH. Successful completion of these Aims will elucidate the contribution(s) of these new necroptosis pathways to autoimmunity and acute liver inflammation with the long-term goal to identify those autoinflammatory/immune diseases that might benefit from RIPK1 kinase targeted therapy