Role of PISA; a PYHIN Family Member in Intracellular DNA Recognition

描述(申请人提供)：先天免疫系统通过胚系编码的模式识别受体感知非自我核酸。RNA通过Toll样受体(TLR)-3、-7和-8或细胞质RNA解旋酶RIG-I和MDA-5感受。对细胞质DNA的传感器知之甚少，当它被传递到细胞质时，会触发I型干扰素基因转录和caspase-1介导的IL-12的处理。我们已经确定PISA(PYHIN蛋白刺激ASC)是四种人类PYHIN蛋白之一，是胞浆DNA的受体。PISA的HIN200结构域与DNA结合，而PYD结构域(但不是PYHIN1-3的结构域)与接头分子ASC结合，激活NF-：B和caspase-1。PISA的敲除下调了caspase-1介导的IL-12反应，伴随着I型干扰素产生的增加。我们的假设是，PISA用于识别微生物胞浆DNA(如细菌或病毒DNA)，PISA通过组装与ASC和caspase-1结合的炎性小体来诱导对细胞质DNA的促炎反应。在这项研究中，我们建议进一步表征PISA在识别病毒和细菌胞浆DNA中的作用。我们还将通过基因捕获技术和/或靶向基因缺失来产生PISA基因敲除小鼠，并建议分析PISA与微生物感染的体内相关性。最后，我们将评估PISA在调节干扰素对细胞质DNA的反应中的作用。阐明胞质DNA识别的分子机制可能会导致微生物感染的新治疗靶点，从而使新的策略能够防止对基因治疗方法的炎症反应。与公共卫生相关：病原体由免疫系统不同细胞区域表达的受体检测。这笔拨款的重点是了解一种胞浆蛋白，我们已经鉴定为PISA(PYHIN蛋白质刺激ASC)是如何通过它们的DNA感知病原体的。阐明PISA激活的分子细节可能导致基于细胞内DNA识别的药物干预疾病的新策略。

DESCRIPTION (provided by applicant): The innate immune system senses non-self nucleic acids via germ-line encoded pattern recognition receptors. RNA is sensed via Toll-like receptor (TLR)-3, -7 and -8 or by the cytoplasmic RNA helicases RIG-I and MDA-5. Little is known about sensors for cytoplasmic DNA, which when delivered to the cytoplasm trigger type I IFN gene transcription and caspase-1-mediated processing of IL-12. We have identified PISA (PYHIN protein stimulating ASC), one of four human PYHIN proteins as a receptor for cytosolic DNA. The HIN200 domain of PISA binds to DNA, while the PYD domain (but not that of PYHIN1-3) associates with the adapter molecule ASC to activate NF-:B and caspase-1. Knockdown of PISA downregulates caspase-1-mediated IL-12 responses, with concomitant increases in type I interferon production. Our hypothesis is that PISA serves to recognize microbial cytosolic DNA (such as bacterial or viral DNA) and that PISA induces pro-inflammatory responses towards cytoplasmic DNA via the assembly of an inflammasome that engages ASC and caspase-1. In this study, we propose to further characterize the role of PISA in the recognition of viral and bacterial cytosolic DNA. We will also generate a PISA knock-out mouse by gene-trap technology and/or targeted gene deletion and propose to analyze the in vivo relevance of PISA for microbial infections. Finally, we will assess the role of PISA for the modulation of interferon responses towards cytoplasmic DNA. Elucidation of the molecular mechanisms of cytosolic DNA recognition could lead to novel therapeutic targets for microbial infections and thereby enable novel strategies that could prevent inflammatory reactions to gene therapy approaches. Public Health Relevance: Pathogens are detected by receptors expressed in different areas of cells of the immune system. This grant is focused on understanding how a cytosolic protein, which we have identified called PISA (PYHIN protein stimulating ASC) senses pathogens by virtue of their DNA. Elucidation of molecular details of PISA activation could lead to novel strategies of pharmacological interference for diseases based on intracellular DNA recognition.