STING-dependent activation of Natural Killer cells by viral and tumor DNA

描述（由申请人提供）：识别宿主细胞细胞质中的外源DNA对于启动对DNA病毒的免疫反应至关重要。病毒DNA的识别激活了许多信号通路，包括诱导强效抗病毒细胞因子，如I型干扰素。虽然细胞质dna感应途径进化为检测病毒，但我们最近的初步数据也表明，dna依赖性的STING激活也可能在肿瘤反应中起关键作用。在这里，我们提出研究细胞质DNA的识别如何导致自然杀伤（NK）细胞的抗肿瘤和抗病毒反应的激活。我们的总体假设是，细胞质检测DNA是NK细胞先天识别病毒和肿瘤的共同机制。我们描述了三个目标，将阐明cGAS-STING途径如何启动抗病毒和抗肿瘤反应。这些目标代表了Raulet实验室（拥有NK细胞和NKG2D激活受体的专业知识）和顾问Russell Vance实验室（拥有细胞质DNA传感和STING途径的专业知识）之间的密切合作。目的如下：(1)确定cGAS-STING通路在体内如何激活NK细胞和其他免疫细胞。我们的初步数据表明，HSV-1或合成的2'3'-cGAMP与STING结合可以激活NK细胞和其他体内免疫细胞。STING激活导致I型IFN、stat6依赖性趋化因子的产生和NKG2D配体的激活。将这些途径与NK激活联系起来的机制，以及所涉及的细胞类型，将在体内使用各种基因敲除小鼠来确定。(2)确定肿瘤的先天免疫控制是否需要cGAS-STING信号。携带E¿-myc转基因的小鼠会患上B细胞淋巴瘤，将与缺乏cGAS-STING信号通路的小鼠杂交。我们将在该模型中确定cGAS-STING的缺失是否会加速肿瘤的发生。我们假设cGAS-STING信号是肿瘤细胞所需的，也可能是激活NK细胞受体（如NKG2D和DNAM1）的配体APC表达所必需的。(3)确定DNA损伤导致胞质DNA积累的机制。我们的初步数据表明，经历DNA损伤或正在进行转化的细胞表现出主要来自内源性逆转录病毒/元件的细胞质DNA的增加和异常积累。在本目的中，我们将评估细胞内DNA的增加是否由于逆转录病毒/元件RNA表达的增加，和/或细胞内逆转录酶活性的增加，和/或细胞内DNA外切酶（如TREX1）活性的降低。

DESCRIPTION (provided by applicant): Recognition of foreign DNA in the cytosol of host cells is critical for the initiation of immune responses to DNA viruses. Recognition of viral DNA activates numerous signaling pathways, including the induction of potent antiviral cytokines such as type I interferons. Although the cytosolic DNA-sensing pathway evolved to detect viruses, our recent preliminary data also suggest that DNA-dependent activation of STING may also be critical in the response to tumors. Here we propose to investigate how recognition of cytosolic DNA leads to activation of anti-tumor and anti-viral responses by Natural Killer (NK) cells. Our overall hypothesis is that cytosolic detection of DNA is a common mechanism underlying innate recognition of viruses and tumors by NK cells. We describe three Aims that will illuminate how the cGAS-STING pathway initiates anti-viral and anti-tumor responses. These Aims represent a close collaboration between the Raulet Lab, which has expertise in NK cells and the NKG2D activating receptor, and the lab of consultant Russell Vance, which has expertise in cytosolic DNA sensing and the STING pathway. The Aims are as follows: (1) Determine how the cGAS-STING pathway activates NK cells and other immune cells in vivo. Our preliminary data show that STING engagement by HSV-1 or synthetic 2'3'-cGAMP activates NK cells and other immune cells in vivo. STING activation leads to production of type I IFN, STAT6-dependent chemokines, and activation of NKG2D ligands. The mechanisms linking these pathways to NK activation, and the cell types involved, will be determined in vivo using various knockout mice. (2) Determine if cGAS-STING signaling is required for innate immune control of tumors. Mice harboring an E¿-myc transgene, which develop B cell lymphomas, will be crossed to mice deficient in the cGAS-STING signaling pathway. We will determine whether loss of cGAS-STING accelerates tumorigenesis in this model. We hypothesize that cGAS-STING signaling is required for tumor cell and possibly APC expression of ligands that activate NK cell receptors (e.g., NKG2D and DNAM1). (3) Determine the mechanisms by which DNA damage leads to accumulation of cytosolic DNA. Our preliminary data indicate that cells experiencing DNA damage or undergoing transformation exhibit increased and aberrant accumulation of cytosolic DNA derived largely from endogenous retroviruses/elements. In this Aim we will assess whether increased cytosolic DNA is due to increased expression of retrovirus/element RNA, and/or increased cytosolic reverse transcriptase activity, and/or decreased activity of cytosolic DNA exonucleases (e.g., TREX1).