Role of RNA-binding protein in immune evasion of Mtb in macrophages

项目总结/摘要 结核病（TB）仍然是一个主要的全球健康问题，约有900万新发病例，近1500万人死于结核病。 全世界每年有100万人死于结核病。结核分枝杆菌（Mtb）感染持续存在， 当宿主免疫反应被抑制或逃避时，巨噬细胞会产生免疫应答。然而，结核病免疫机制 逃避仍然是不完整的。更好地理解这些机制将有助于开发主机导向的 治疗（HDT），特别是对致命的耐药结核病。我们的初步数据表明， 三曲脯氨酸（TTP）蛋白有助于逃避，可以成为宿主导向的TB治疗的靶点。TTP是RNA 在转录后水平参与炎症反应调节的β-结合蛋白。TTP结合 在3 '非翻译区（3'UTR）内的腺嘌呤-尿苷富集元件（ARE），导致 我们和其他人已经证明，编码几种对Mtb清除很重要的细胞因子的mRNA是一种免疫抑制剂。 维持体内平衡重要调节器。然而，TTP是否影响Mtb复制或再活化， 仍然未知。我们首次在结核分枝杆菌感染的野生型小鼠肺中发现了显著的TTP诱导 在感染分枝杆菌的巨噬细胞中，缺失TTP导致分枝杆菌 巨噬细胞的生长。此外，用活的Mtb攻击的TTP缺陷小鼠中的细菌显著减少。 肺比感染野生型同窝仔。酪样肉芽肿组织中TTP mRNA水平显著高于对照组， 肺结核患者的肺实质较正常肺实质明显。结核病的复发和传播涉及干酪性 形成空洞并释放结核分枝杆菌的肉芽肿。此外，我们的数据首次表明，抑制mTOR 雷帕霉素的信号传导几乎完全消除了Mtb感染的巨噬细胞中的TTP诱导。基于我们 研究发现，我们提出了mTOR/TTP轴在结核免疫逃避中的新作用：毒力Mtb激活mTOR 诱导TTP的途径，导致抗TB免疫、Mtb复制和原发性和/或潜伏性抑制 再活化疾病TTP可作为逆转Mtb免疫逃避的靶点。我们提出两个目标， 测试这个新的假设：目标1。确定TTP在巨噬细胞中对mTOR介导的Mtb生长的作用 以及靶向TTP对Mtb生长的影响，用于潜在的宿主定向治疗。目标2.探讨其分子 Mtb在巨噬细胞中诱导TTP的机制TTP诱导机制的确定将不会 这不仅提高了我们对结核分枝杆菌逃避机制的理解，而且还提供了可用于 开发针对宿主的疗法，打破结核病逃避。这些探索性和机制性研究的结果将 更好地了解结核病免疫逃避机制，并可能导致TTP靶向治疗， 尤其是耐药结核病。

Project Summary/Abstract Tuberculosis (TB) remains a leading global health problem with about 9 million new cases and nearly 1.5 million TB-related deaths worldwide each year. Mycobacterium tuberculosis (Mtb) infection persists in macrophages when the host immune response is suppressed or evaded. However, mechanisms of TB immune evasion remain incomplete. Better understanding the mechanisms will facilitate development of host-directed therapy (HDT), especially against deadly drug-resistant Mtb. Our preliminary data indicate that an RNA-binding protein, tristetraprolin (TTP) facilitates evasion and can be a target for host-directed TB therapy. TTP is an RNA -binding protein involved in the regulation of inflammatory responses at the post-transcriptional level. TTP binds to adenine-uridine-rich elements (AREs) within the 3’ untranslated region (3'UTR) causing destabilization of mRNAs encoding several cytokines important for Mtb clearance, and has been shown by us and others to be an important regulator maintaining homeostasis. Whether TTP affects Mtb replication or reactivation, however, remains unknown. We found for the first time a significant TTP induction in lungs of Mtb-infected wild type mice and in macrophages infected with mycobacteria, and deleting TTP resulted in suppression of mycobacterial growth in macrophages. Further, mice deficient in TTP challenged with live Mtb had significantly less bacteria in lungs than infected wild type littermates. TTP mRNA levels were significantly higher in caseous granulomas of TB patients than in normal lung parenchyma. Reactivation and transmission of TB involve developing caseous granulomas that cavitate and release Mtb. In addition, our data show for the first time that inhibition of mTOR signaling by rapamycin almost completely abolishes TTP induction in Mtb-infected macrophages. Based on our findings, we propose a novel role for mTOR/TTP axis in TB immune evasion: Virulent Mtb activates mTOR pathway that induces TTP, resulting in suppression of anti-TB immunity, Mtb replicates and primary and/or latent reactivation diseases. TTP may serve as a target to reverse immune evasion of Mtb. We propose two aims to test this novel hypothesis: Aim 1. Determine the role of TTP in macrophages for mTOR-mediated Mtb growth and the effects of targeting TTP on Mtb growth for potential host-directed therapy. Aim 2. Explore the molecular mechanisms of TTP induction by Mtb in macrophages. Identification of the mechanisms of TTP induction will not only enhance our understanding of Mtb evasion mechanisms, but also provide targets that can be used to develop host-directed therapy breaking TB evasion. Results of these exploratory and mechanistic studies will provide a better understanding of TB immune evasion mechanisms and potentially lead to TTP-targeted therapy, especially with drug-resistant Mtb.