The piRNA binding protein Miwi2 promotes cytokine expression during bacterial pneumonia

 描述（由申请方提供）：急性下呼吸道感染是全球最大的疾病负担，自抗生素发现以来，死亡率几乎保持不变。有效的免疫反应需要严格调节细胞因子和趋化因子的产生，这些细胞因子和趋化因子可以促进最大限度的细菌清除，同时最大限度地减少组织损伤。需要更好地了解肺中细胞因子的调节，因为考虑到抗生素耐药性的上升，未来针对免疫反应的治疗可能是必不可少的。作为未来治疗的基础，我们的实验室有兴趣确定新的调节因子的制定。一个主要的监管机构 在基因表达中，PIWI蛋白与哺乳动物生殖系中的PIWI相互作用RNA（piRNA）结合，并起抑制古老的逆转录病毒元件的作用。小鼠中三种PIWI蛋白（MIWI、MILI或MIWI 2）中任何一种的消融都会导致由于LINE元件的异常表达而导致的精子发生缺陷。虽然已经描述了PIWI蛋白的生殖细胞功能，但几乎不知道它们在体细胞中的潜在作用。令我们惊讶的是，对分选的肺泡上皮细胞的微阵列分析和qRT-PCR证明了在细菌性肺炎期间在肺中诱导单一PIWI蛋白Miwi 2。Miwi 2的体外和体内消耗均表明其在相关炎症条件下促进细胞因子和趋化因子表达。据我们所知，这是Miwi 2的体细胞功能的第一次证明。鉴于这些调查结果，仍然存在几个重要问题。我们将检验中心假设，即在细菌性肺炎期间，piRNA结合蛋白Miwi 2从其已知的作为古老病毒元件的生殖细胞特异性阻遏物的功能被重新利用，并在肺上皮细胞中被诱导以促进细胞因子表达和免疫防御。在具体目标1中，我们将使用一种荧光流式细胞术策略来识别特定的肺泡 上皮细胞群，其中Miwi 2在肺炎期间被诱导。在特定目标2中，我们开始阐明Miwi 2依赖性细胞因子诱导的分子机制。在具体目标3中，我们检验了以下假设：细胞因子和趋化因子的Miwi 2依赖性表达对于细菌性肺炎的临床相关模型中的宿主防御是必需的。这些研究的结果将阐明皮尔纳结合蛋白的未被描述的体细胞功能，以及增强我们对细菌性肺炎期间细胞因子调节的理解。由于皮尔纳结合蛋白诱导与某些癌症相关，我们预计这些数据将超出我们目前对先天免疫的关注。希望这里获得的结果将作为未来免疫调节疗法的基础，改善肺炎患者的预后。

 DESCRIPTION (provided by applicant): Acute lower respiratory tract infections represent the greatest burden of disease worldwide, with mortality rates that have remained virtually unchanged since the discovery of antibiotics. The effective immune response requires the tightly regulated production of cytokines and chemokines that can facilitate maximal bacterial clearance, while minimizing tissue damage. A better understanding of cytokine regulation in the lung is needed, as future therapeutics targeting the immune response may be an essential given the rising rates of antibiotic resistance. As a foundation for future therapeutics, our laboratory is interested in identifying novel regulators of cytokine elaboration. A major regulator of gene expression, PIWI proteins associate with PIWI- interacting RNAs (piRNAs) in the mammalian germ line, and function to repress ancient retroviral elements. Ablation of any of the three PIWI proteins in mice (MIWI, MILI, or MIWI2) results in defects in spermatogenesis due to aberrant expression of LINE elements. While the germ cell functions of PIWI proteins have been described, next to nothing is known about their potential role(s) in somatic cells. To our surprise microarray analysis and qRT-PCR of sorted alveolar epithelial cells demonstrated that a single PIWI protein, Miwi2 is induced in the lung during bacterial pneumonia. Both in vitro and in vivo depletion of Miwi2 indicates that it acts to promote cytokine and chemokine expression under relevant inflammatory conditions. To our knowledge, this is the first demonstration of a somatic cell function for Miwi2. Given these findings, several important questions still remain. We will test the central hypothesis that during bacterial pneumonia, the piRNA-binding protein Miwi2 is repurposed from its known function as a germ cell specific repressor of ancient viral elements and is induced in pulmonary epithelial cells to promote cytokine expression and immune defense. In Specific Aim 1 we will use a multicolor FACS strategy to identify the specific alveolar epithelial cell population where Miwi2 is induced during pneumonia. In Specific Aim 2 we begin to elucidate the molecular mechanisms of Miwi2 dependent cytokine induction. In Specific Aim 3 we test the hypothesis that Miwi2 dependent expression of cytokines and chemokines are necessary for host defense in a clinically relevant model of bacterial pneumonia. Results of these studies will shed light on the under-described somatic functions of piRNA binding proteins, as well as enhance our understanding of cytokine regulation during bacterial pneumonia. As piRNA binding protein induction has been associated with certain cancers, we anticipate that these data will extend beyond our current focus of innate immunity. Hopefully, the results gained here will serve as a basis for future immunomodulatory therapeutics that improves the outcomes of patients with pneumonia.