Translation initiation factors driving persistence of Toxoplasma gondii bradyzoites in neurons

驱动弓形虫缓殖子在神经元中持续存在的翻译起始因子

• 批准号: 10556561
• 负责人: William J Sullivan
• 金额: $ 57.38万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-12-08 至 2027-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10556561
• 关键词: Acute; Address; Animals; Area; Automobile Driving; Binding; Brain; Cells; Chronic; Clinical; Collaborations; Collection; Complex; Cyst; Data; Development; Eukaryotic Initiation Factor-2; Goals; Human; Immune; Immunosuppression; Individual; Infection; Initiator Codon; Knowledge; Laboratories; Life; Messenger RNA; Modeling; Myocardium; Neurologic; Neurons; Nucleic Acid Regulatory Sequences; Open Reading Frames; Opportunistic Infections; Parasites; Patients; Peptide Initiation Factors; Pharmacotherapy; Phosphorylation; Phosphotransferases; Physiological; Population; Proliferating; Protein Biosynthesis; Protein Synthesis Induction; RNA; RNA Helicase; Recurrence; Regulatory Element; Ribosomes; Risk; Role; Scanning; Signal Transduction; Skeletal Muscle; Stress; Structure; Symptoms; System; Therapeutic Intervention; Tissues; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Transfer RNA; Translating; Translation Initiation; Translations; Work; combat; enhancer-binding protein AP-2; helicase; innovation; insight; mRNA Translation; mRNA capping; new therapeutic target; novel; novel therapeutics; prevent; programs; recruit; response; side effect; stress management; stress reduction; transcription factor; translatome

PROJECT SUMMARY The protozoan parasite Toxoplasma gondii can cause recurring opportunistic infections due to its ability to persist as a latent form (bradyzoite) within patients. There is no treatment that targets bradyzoite cysts, which reconvert into the destructive proliferative stage (tachyzoites) in the immune compromised. Patients suffering from reactivated toxoplasmosis frequently present with life-threatening neurological problems, underscoring the significance of bradyzoite cysts in the brain. A better understanding of the mechanisms that drive the development of bradyzoites in neurons is necessary to devise new therapies that prevent their formation and persistence. To address this need, we developed a novel model of spontaneous tissue cyst formation in neurons using Lund human mesencephalic (LUHMES) cells. We will use this innovative system to determine the mechanisms underlying the changes in protein synthesis that are required for bradyzoite formation. Based on our previous collaborative work, we hypothesize that translation initiation factors coordinate changes in 5’-leader sequences of key mRNAs, resulting in changes in protein synthesis that induce conversion to bradyzoites. Translation begins with the binding of an eIF4F complex to the 5’-cap, which then recruits eIF2, which carries Met-tRNA. We previously showed TgIF2 is phosphorylated during bradyzoite conversion, which lowers its abundance and can alter start codon selection. Aim 1 will determine how TgIF2 becomes phosphorylated and which mRNAs are preferentially translated during spontaneous bradyzoite conversion in neurons. Our RiboSeq approach will reveal areas within 5’-leaders that regulate translation, such as upstream open reading frames (uORFs) or secondary structures, the latter of which will be identified by generating the first RNA “structurome” for Toxoplasma in Aim 2. As these structures are resolved by helicase activity of eIF4F, Aim 2 will also delineate the functions of the multiple TgIF4F complexes we have uncovered in the parasites. Together, these studies will determine how cellular signals coordinate translation initiation factors to reprogram the translatome to trigger the spontaneous formation of bradyzoites in human neurons. Completion of this study will have a sustained high impact on the field by providing significant new insights into the complex mechanisms that Toxoplasma uses to persists in its host, which will reveal novel points for therapeutic intervention.

项目总结 原生动物寄生虫弓形虫可以由于其持续存在的能力而导致反复的机会性感染。 作为患者体内的一种潜伏形式(缓殖子)。目前还没有针对慢体包囊的治疗方法，这些包囊会重新转化。 进入破坏性增殖阶段(速殖子)的免疫受损。患者患有 再激活的弓形虫病经常出现危及生命的神经问题，突显出 脑部慢形虫囊肿的临床意义。更好地了解推动经济增长的机制 在神经元中发育缓殖子是必要的，以设计新的治疗方法来防止它们的形成和 坚持不懈。为了满足这一需求，我们开发了一种新的神经元自发组织囊形成模型。 使用隆德人中脑(LUHMES)细胞。我们将使用这个创新的系统来确定 缓殖子形成所需的蛋白质合成变化的潜在机制。基于 在我们之前的合作工作中，我们假设翻译启动因素协调了 关键mRNAs的5‘-前导序列，导致蛋白质合成的变化，从而诱导转化为 缓虫体。翻译开始于将eIF4F复合体与5‘-帽结合，然后招募eIF2， 携带Met-tRNA。我们之前展示了TgIF2在缓殖子转换过程中被磷酸化，这是 降低其丰度，并可能改变起始密码子的选择。目标1将决定TgIF2如何成为 在缓殖子自发转换过程中，哪些mRNAs优先被翻译 神经元。我们的RiboSeq方法将揭示5‘-领导者中规范翻译的区域，例如上游 开放阅读框架(UORF)或二级结构，后者将通过生成第一个 Aim 2中弓形虫的RNA“结构”。由于这些结构是通过eIF4F的解旋酶活性来解析的，所以Aim 2还将描述我们在寄生虫中发现的多个TgIF4F复合体的功能。 总之，这些研究将确定细胞信号如何协调翻译启动因子来重新编程 在人类神经元中触发缓殖子自发形成的翻译组。完成这项研究 通过提供对复杂机制的重大新见解，将对该领域产生持续的高度影响 弓形虫用来在宿主体内持续存在的物质，这将揭示治疗干预的新观点。