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.

项目概要 原生动物寄生虫弓形虫由于其持续存在的能力，可引起反复的机会性感染 作为患者体内的潜伏形式（缓殖子），没有针对缓殖子包囊的治疗方法，缓殖子包囊会重新转变。 免疫受损的患者进入破坏性增殖阶段（速殖子）。 重新激活的弓形体病经常出现危及生命的神经系统问题，这强调了 缓殖子囊肿在大脑中的重要性更好地理解驱动机制。 神经元中缓殖子的发育对于设计新的疗法来阻止其形成和 为了满足这一需求，我们开发了一种神经元自发组织囊肿形成的新模型。 我们将使用隆德人类中脑 (LUHMES) 细胞来确定 缓殖子形成所需的蛋白质合成变化的机制。 我们之前的合作工作中，我们勇敢地认为翻译启动因素协调了 关键 mRNA 的 5’-前导序列，导致蛋白质合成发生变化，从而诱导转化为 翻译从 eIF4F 复合物与 5’-cap 的结合开始，然后招募 eIF2， 我们之前表明 TgIF2 在缓殖子转化过程中被磷酸化。 降低其丰度并可以改变起始密码子选择 目标 1 将决定 TgIF2 如何变化。 磷酸化以及哪些 mRNA 在自发缓殖子转化过程中优先翻译 我们的 RiboSeq 方法将揭示 5’-前导区域内调节翻译的区域，例如上游。 开放阅读框（uORF）或二级结构，后者将通过生成第一个结构来识别 Aim 2 中弓形虫的 RNA“结构组”。由于这些结构是通过 eIF4F 的解旋酶活性解析的，Aim 图 2 还将描述我们在寄生虫中发现的多个 TgIF4F 复合物的功能。 这些研究将共同​​确定细胞信号如何协调翻译起始因子以进行重编程 翻译组触发人类神经元中缓殖子的自发形成 完成这项研究。 通过提供对复杂机制的重要新见解，将对该领域产生持续的重大影响 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点。