The translation apparatus of Leishmania: from basic analysis to pursuit of novel

利什曼原虫的翻译机构：从基础分析到小说追求

基本信息

批准号：
9297201
负责人：
GERHARD WAGNER
金额：
$ 34.87万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-08-06 至 2019-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9297201
关键词：
Adverse effects Affect Appearance Back Binding Biochemical Biological Assay Biology Bite Chemicals Chemistry Clinical Code Complex Country Crystallization Data Development Disease Drug Targeting Effectiveness Environment Epitopes Eukaryota Family Female Fluorescence Fluorescence Polarization Genes Genome Geography Goals Human In Vitro Infection Institutes Interruption Intravenous Leishmania Leishmania major Leishmaniasis Length Life Ligands Light Malignant Neoplasms Mammalian Cell Measurement Measures Messenger RNA Molecular Molecular Biology Monitor NMR Spectroscopy Organism Outcome Parasite resistance Parasites Parasitology Peptide Initiation Factors Peptides Pharmacotherapy Phlebotominae Plasmodium falciparum Protein Biosynthesis Proteins Proteolysis RNA Cap-Binding Proteins Recruitment Activity Relaxation Research Research Proposals Residual state Resistance Resolution Ribosomes Roentgen Rays Role Route Scaffolding Protein Sequence Homology Site Stress Structure Syndrome System Time Translation Initiation Translations Trypanosoma brucei brucei Trypanosoma cruzi Trypanosomatina X-Ray Crystallography base biophysical techniques effective therapy experimental study flexibility high throughput screening inhibitor/antagonist medical attention medical schools member neglect new therapeutic target novel novel therapeutics pathogen protein complex protein protein interaction public health relevance screening small molecule inhibitor tool

项目摘要

DESCRIPTION (provided by applicant): Protozoan parasites that belong to the genus Leishmania from the Trypanosomadae family, causes Leishmaniasis in humans, which affects 350 million people in 88 countries worldwide. The parasites are principally transmitted to humans following the bite of female phlebotomine sandflies, which carry the infectious stage of the organism. Leishmaniasis comprise three major clinical syndromes principally determined by the species of infecting parasites, which vary geographically. Few therapies are currently available, but strong side effects and parasite resistance limit their effectiveness. The recent advancements in determining several trypanosomatid genomes provided an invaluable tool to gather information regarding the molecular and biochemical mechanisms used by these organisms. However, it has proven difficult to correctly assign specific functions to gene coding sequences because the sequence homology between these parasites and their higher eukaryotic counterparts is extremely low. Until recently, little effort was invested in the Leishmania cap-binding complexes in the human stage, which are required to initiate protein synthesis in parasites. In this proposal, we focus on three translation initiation factors from Leishmania major in the human stage: LeishIF4E-1, Leish4E-IP and LeishIF3. While LeishIF4E-1 has emerged as the crucial cap-binding protein, it is hypothesized that Leish4E-IP and LeishIF3 are, respectively, a regulator of translation initiation and a scaffold protein responsibl for recruiting the small ribosomal subunit at the 5' end of messenger RNAs. To investigate whether this is the case, we will apply a combination of biophysical techniques, including NMR spectroscopy and X-ray crystallography, and molecular biology approaches to study the structure/function of these factors in vitro and within parasites. Once we determine indispensable interactions that promote translation initiation in the parasite, we will explore these potential targets in high-throughput (HTP) small molecule inhibitor screens. This proposal will allow us to highlight similarities and differences between translation initiation in human cels and Leishmania parasites, and validate the Leishmania translation machinery as a potential drug target. This concept is backed by data obtained previously in the Wagner lab where a crucial interaction between two human translation initiation factors (eIF4E/eIF4G) has been targeted in a HTP screen and led to the discovery of compounds with anti-cancer activity. The research will pursue three specific aims: 1. Determine the structure of the LeishIF4E-1 complex with its ligand cap-4 and Leish4E-IP. 2. Characterize the LeishIF3 recruitment to the Leishmania cap-binding complex. 3. Develop assays for HTP screens in search of Leishmania inhibitory compounds.

描述（由申请人提供）：属于锥虫科利什曼原虫属的原生动物寄生虫会引起人类利什曼病，影响全球 88 个国家的 3.5 亿人。这种寄生虫主要通过雌性白蛉叮咬后传播给人类，雌性白蛉携带该生物体的感染阶段。利什曼病包括三种主要的临床综合征，主要由感染寄生虫的种类决定，而这些种类因地理位置而异。目前可用的疗法很少，但强烈的副作用和寄生虫耐药性限制了其有效性。最近在确定几种锥虫基因组方面取得的进展为收集有关这些生物体所使用的分子和生化机制的信息提供了宝贵的工具。然而，事实证明很难正确地将特定功能分配给基因编码序列，因为这些寄生虫与其高等真核对应物之间的序列同源性极低。直到最近，人们对人类阶段的利什曼原虫帽结合复合物投入很少，而这种复合物是启动寄生虫蛋白质合成所必需的。在本提案中，我们重点关注人类阶段主要利什曼原虫的三个翻译起始因子：LeishIF4E-1、Leish4E-IP 和 LeishIF3。虽然 LeishIF4E-1 已成为关键的帽结合蛋白，但推测 Leish4E-IP 和 LeishIF3 分别是翻译起始的调节因子和负责招募信使 5' 端核糖体小亚基的支架蛋白RNA。为了调查情况是否如此，我们将应用生物物理技术（包括核磁共振波谱学和 X 射线晶体学）和分子生物学方法的组合来研究体外和寄生虫内这些因子的结构/功能。一旦我们确定了促进寄生虫翻译起始的不可或缺的相互作用，我们将在高通量（HTP）小分子抑制剂筛选中探索这些潜在靶标。该提案将使我们能够强调人类细胞和利什曼原虫寄生虫翻译起始之间的相似性和差异，并验证利什曼原虫翻译机制作为潜在的药物靶点。这一概念得到了瓦格纳实验室先前获得的数据的支持，该实验室在 HTP 筛选中针对两个人类翻译起始因子 (eIF4E/eIF4G) 之间的关键相互作用，并导致了具有抗癌活性的化合物的发现。该研究将追求三个具体目标： 1. 确定 LeishIF4E-1 复合物及其配体 cap-4 和 Leish4E-IP 的结构。 2. 表征 LeishIF3 募集到利什曼原虫帽结合复合物的特征。 3. 开发 HTP 筛选分析方法以寻找利什曼原虫抑制化合物。

项目成果

期刊论文数量（3）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Structural basis for LeishIF4E-1 modulation by an interacting protein in the human parasite Leishmania major.

DOI：
10.1093/nar/gky194
发表时间：
2018-04-20
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Meleppattu S;Arthanari H;Zinoviev A;Boeszoermenyi A;Wagner G;Shapira M;Léger-Abraham M
通讯作者：
Léger-Abraham M

The eIF3 complex of Leishmania-subunit composition and mode of recruitment to different cap-binding complexes.

利什曼原虫亚基组成的 eIF3 复合物以及招募不同帽结合复合物的模式。

DOI：
10.1093/nar/gkv564
发表时间：
2015
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Meleppattu,Shimi;Kamus-Elimeleh,Dikla;Zinoviev,Alexandra;Cohen-Mor,Shahar;Orr,Irit;Shapira,Michal
通讯作者：
Shapira,Michal

A newly identified Leishmania IF4E-interacting protein, Leish4E-IP2, modulates the activity of cap-binding protein paralogs.

新鉴定的利什曼原虫 IF4E 相互作用蛋白 Leish4E-IP2 可调节帽结合蛋白旁系同源物的活性。

DOI：
10.1093/nar/gkaa173
发表时间：
2020
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Tupperwar,Nitin;Meleppattu,Shimi;Shrivastava,Rohit;Baron,Nofar;Gilad,Ayelet;Wagner,Gerhard;Léger-Abraham,Mélissa;Shapira,Michal
通讯作者：
Shapira,Michal