RAPs-mediated post-transcriptional control in Apicomplexan parasites

RAP 介导的顶复门寄生虫转录后控制

• 批准号: 9788270
• 负责人: Karine Gaelle Le Roch
• 金额: $ 56.06万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-19 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9788270
• 关键词: Address; Antimalarials; Antiparasitic Agents; Aptamer Technology; Binding Proteins; Biochemical; Biogenesis; Bioinformatics; Biology; CRISPR/Cas technology; Cell physiology; Cessation of life; Characteristics; Code; Communicable Diseases; Computer Analysis; Data; Development; Drug Targeting; Engineering; Epitopes; Erythrocytes; Eukaryota; Family; Future; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genomics; Goals; Health; High-Throughput Nucleotide Sequencing; Human; In Situ; Knowledge; Lead; Life Cycle Stages; Maintenance; Malaria; Mass Spectrum Analysis; Mediating; Messenger RNA; Molecular; Morbidity - disease rate; Nuclear; Nuclear Export; Organelles; Organism; Parasites; Pathway interactions; Peptide Elongation Factor 2; Phase; Phenotype; Plasmodium; Plasmodium falciparum; Play; Post-Transcriptional Regulation; Protein Binding Domain; Protein Biosynthesis; Protein Family; Proteins; Proteomics; Publishing; RNA; RNA Recognition Motif; RNA Splicing; RNA-Binding Proteins; Research; Resistance; Ribonucleoproteins; Role; Technology; Therapeutic; Therapeutic Intervention; Time; Transcript; Translations; Vaccines; Validation; Virulence; base; comparative; crosslinking and immunoprecipitation sequencing; design; drug discovery; experimental study; genome-wide; human pathogen; improved; insight; mRNA Stability; mortality; multidisciplinary; new therapeutic target; novel; novel therapeutic intervention; novel therapeutics; pre-clinical; programs; protein expression; small molecule inhibitor; transcriptome; transcriptome sequencing; zygote

Project Summary/Abstract In the malaria parasite, gene regulation is heavily dependent on mechanisms acting at the post- transcriptional and translational levels. More importantly, Plasmodium possesses many atypical characteristics in both pathways when compared to its human host. These unusual features hence extend our therapeutic window against this deadly parasite, yet these characteristics have been poorly explored at the molecular level. The main goal of this project is to characterize the role of newly identified apicomplexan-specific RNA- binding proteins {RAPs) in parasite development and virulence, and to validate their potential as novel drug targets. The fact that Plasmodium parasites modulate part of his gene expression at the post-transcriptional level was inspired by many early studies that demonstrated that the global proteomic profiles often lagged behind the transcriptional profiles. Our newly published large-scale bioinformatics reinforce this conjecture as we determined that at least 18% of all coding-gene are predicted to be potential RNA binding domain-containing protein (RBP). Furthermore, many of these parasite RBPs were experimentally shown to interact in situ with mRNAs. In all eukaryotic organisms, RBPs are essential to regulate mRNA processing at multiple levels including splicing, transport, mRNA stability and turnover, as well as mRNA localization and translational efficiency. In the human malaria parasite, very few RBPs have been characterized. We propose to address this knowledge gap by examining the function of the parasite specific RNA-binding domain proteins abundant in Apicomplexans, or RAPs. More specifically, we will use state-of-the-art genomics, molecular, and cellular approaches to determine the role of the 21 identified RAPs in parasite development and survival. In Aim 1, we will characterize the essentiality, subcellular localization and phenotypes of RAPs in P. falciparum across different developmental stage using novel CRISPR/Cas9 approaches. In Aim 2, we will define the global RAP- related ribonucleoprotein interaction network across various parasite developmental stages by developing high-throughput sequencing technologies and pull-down strategies followed by mass spectrometry analysis. Our complementary approaches will not only identify key specific RBPs contributing to parasite development, but also uncover unique post-transcriptional networks in a eukaryotic human pathogen. By providing fundamental insights into mechanisms regulating translation in Plasmodium, this project will improve our ability to design new drugs and novel lines of defense against malaria and many other infectious disease agents.

项目总结/摘要 在疟疾寄生虫中，基因调控在很大程度上依赖于作用于 转录后和翻译水平。更重要的是，疟原虫拥有许多 与人类宿主相比，这两种途径都具有非典型特征。这些不寻常 因此，这些功能扩展了我们对这种致命寄生虫的治疗窗口，但这些 这些特征在分子水平上探索得很差。该项目的主要目标是 为了表征新鉴定的顶复体特异性RNA结合蛋白（RAP）在 寄生虫的发育和毒力，并验证其作为新型药物靶点的潜力。 疟原虫在转录后水平调节部分基因表达， 水平的灵感来自于许多早期的研究，这些研究表明，全球蛋白质组学概况往往 落后于转录谱。我们新发表的大规模生物信息学 我们确定至少有18%的编码基因是预测的， 是潜在的RNA结合域蛋白（RBP）。此外，许多寄生虫 实验表明RBP与mRNA原位相互作用。在所有真核生物中， RBP对于在多个水平上调节mRNA加工是必不可少的，包括剪接， 转运，mRNA稳定性和周转，以及mRNA定位和翻译 效率在人类疟疾寄生虫中，很少有限制性商业惯例被描述。我们建议 通过检查寄生虫特异性RNA结合的功能来解决这一知识空白 在顶复门中丰富的结构域蛋白，或RAP。更具体地说，我们将使用 最先进的基因组学，分子和细胞方法，以确定21 确定了寄生虫发育和存活的RAP。在目标1中，我们将描述 恶性疟原虫RAPs的重要性、亚细胞定位和表型 不同的发育阶段使用新的CRISPR/Cas9方法。在目标2中，我们将定义 跨多种寄生虫的RAP相关核糖核蛋白相互作用网络 通过开发高通量测序技术和下拉技术， 策略，然后进行质谱分析。我们的互补方法不仅将 确定有助于寄生虫发展的关键特定RBP，但也发现独特的 真核人类病原体中的转录后网络。通过提供 对疟原虫翻译调节机制的基本见解，该项目将 提高我们设计新药物和新防线的能力， 传染病病原体。