Ribosome Profiling in Plasmodium falciparum

恶性疟原虫核糖体分析

基本信息

批准号：
8661701
负责人：
Peter John Myler
金额：
$ 23万
依托单位：
SEATTLE BIOMEDICAL RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-05-15 至 2016-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8661701
关键词：
Attention Binding Bioinformatics Biological Assay Blood Code Complex Couples Coupling Culicidae Data Development Developmental Process Disease Drug resistance Erythrocytes Exons Future Gene Expression Generations Genes Genetic Translation Genome Goals Heat-Shock Response Human Individual Infection Ingestion Initiator Codon Introns Knowledge Left Leishmania Libraries Life Life Cycle Stages Liver Location Malaria Measures Mediating Messenger RNA Military Personnel Morbidity - disease rate Nutrient Open Reading Frames Organism PAWR protein Pancreatic ribonuclease Parasites Peptide Initiation Factors Pharmaceutical Preparations Phase Phosphorylation Phosphotransferases Plasmodium Plasmodium falciparum Play Pregnancy Proteins Proteome Proteomics RNA RNA library RNA-Binding Proteins Ribosomes Role Saccharomyces cerevisiae Seminal Sexual Development Signal Transduction Specific qualifier value Sporozoites Staging Stress Structure System Technology Testing Time Toxoplasma Transcript Translating Translational Regulation Translational Repression Translations Trypanosoma Trypanosoma brucei brucei Vaccines Variant Virus Diseases Work Yeasts asexual deep sequencing environmental change extracellular fighting genome annotation improved killings mRNA Expression new technology next generation sequencing nuclease programs public health relevance research study response tool transcriptome sequencing transmission process

项目摘要

DESCRIPTION (provided by applicant): The malaria parasite Plasmodium falciparum kills over 650,000 people a year and causes widespread morbidity. Human infection commences with a silent phase in the liver, after which the parasites emerge into the blood to infect red blood cells. There, the parasites develop in an ordered, yet graded, progression from rings to trophozoites to schizonts before escaping as merozites to start another erythrocytic cycle and disease. During the erythrocytic cycle, a structured cycle of mRNA abundance is evident, but the correlation between protein and mRNA levels is, at best, modest. This project explores the hypothesis that translational regulation tunes the proteome in the face of a regimented program for mRNA expression. Recently work has shown that translational controls are likely to be important in the erythrocytic cycle, as well as in the transition from mosquito sporozoites to live stages and in the development of the sexual stages essential to transmission. A corollary of these studies is that gene products that are highly regulated at the level of translation are likel important modulators of developmental change and the parasite's response to environmental stress. The project goals are to globally and quantitatively assess the rate at which each mRNA is actively translated in specific erythrocytic stages by coupling the ability to isolate the speciic "footprints" of mRNAs that are occupied by ribosomes (an indicator of translation) with the depth and breadth of next generation sequencing. Aim 1 will establish the ribosome footprinting technology in P. falciparum using trophozoite stage parasites. It will optimize conditions for nuclease treatment to generate the mRNA footprints protected by ribosomes and for the generation of unbiased libraries from the RNA footprints for Illumina deep sequencing. It will also include maturation of the bioinformatics pipeline to analyze resulting sequence data, adapting existing pipelines that are used for RNA-Seq in P. falciparum and for ribosome footprinting of trypanosomes. Aim 2 will expand into other stages of the asexual erythrocytic cycle, identifying genes that are differentially translated across these pathogenic stages. It will also examine gametocytes, which are poised for an abrupt environmental change upon ingestion by the mosquito. The proposed work will yield both an overview of the extent of translational control in P. falciparum blood stages, as well as a quantification of the translationl control of individual gene products. From such data, additional hypotheses on mechanisms of control and functions of regulated proteins can be generated and tested. The project will also improve genome annotation by determining whether proteins are translated from mRNAs corresponding to individual open reading frames, particularly those annotated as hypothetical proteins and those encoded by non-canonical protein-coding open reading frames, to provide a comprehensive view of the translational landscape of P. falciparum erythrocytic stages.

描述（由申请人提供）：疟原虫恶性疟原虫每年导致超过 650,000 人死亡并导致广泛发病。人类感染从肝脏中的静默期开始，之后寄生虫进入血液中感染红细胞。在那里，寄生虫以有序但分级的方式发育，从环到滋养体再到裂殖体，然后作为裂殖子逃逸，开始另一个红细胞周期和疾病。在红细胞周期中，mRNA 丰度的结构化循环是明显的，但蛋白质和 mRNA 水平之间的相关性充其量是适度的。该项目探讨了这样的假设：面对 mRNA 表达的严格程序，翻译调控会调整蛋白质组。最近的研究表明，翻译控制可能在红细胞周期、从蚊子子孢子到活阶段的转变以及传播所必需的有性阶段的发育中发挥重要作用。这些研究的一个推论是，在翻译水平上受到高度调控的基因产物可能是发育变化和寄生虫对环境应激反应的重要调节剂。该项目的目标是通过将分离核糖体占据的 mRNA 的特定“足迹”（翻译指标）的能力与深度和深度结合起来，全面定量地评估每个 mRNA 在特定红细胞阶段的主动翻译速率。下一代测序的广度。目标 1 将利用滋养体阶段寄生虫在恶性疟原虫中建立核糖体足迹技术。它将优化核酸酶处理的条件，以生成受核糖体保护的 mRNA 足迹，并从 RNA 足迹生成无偏差文库，用于 Illumina 深度测序。它还将包括生物信息学管道的成熟，以分析所得序列数据，调整用于恶性疟原虫RNA测序和锥虫核糖体足迹的现有管道。目标 2 将扩展到无性红细胞周期的其他阶段，识别在这些致病阶段差异翻译的基因。它将还检查配子细胞，配子细胞在被蚊子摄入后会发生突然的环境变化。拟议的工作将产生对恶性疟原虫血液阶段翻译控制程度的概述，以及对单个基因产物的翻译控制的量化。根据这些数据，可以生成并测试有关受调节蛋白质的控制机制和功能的额外假设。该项目还将通过确定蛋白质是否从与各个开放阅读框相对应的mRNA翻译来改进基因组注释，特别是那些注释为假设蛋白质和由非规范蛋白质编码开放阅读框编码的蛋白质，以提供翻译的全面视图。恶性疟原虫红细胞阶段的景观。