Translational partitioning of the SL RNA

SL RNA 的翻译分配

基本信息

批准号：
7847643
负责人：
DAVID A CAMPBELL
金额：
$ 19.25万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-06-01 至 2012-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7847643
关键词：
Binding Binding Proteins Biogenesis Biological Assay Cell Nucleus Cells Cloning Complementary DNA Complex Cytosol Data Discrimination EIF4EL3 gene Eukaryota Exhibits Family Gene Expression Homologous Gene Human Kinetoplastida Lead Leishmania Leishmania major Masks Mass Spectrum Analysis Medical Messenger RNA Methylation Mutate Nature Nuclear Nucleotides Organism Pathway interactions Peptide Initiation Factors Phenotype Play Polyribosomes Population Primer Extension Process Property Protein Biosynthesis Proteins RNA RNA Cap-Binding Proteins RNA Caps RNA Processing RNA Splicing RNA-Binding Proteins Small Nuclear RNA Specificity Spliced Leader RNA Spliced Leader Sequences Structure System Touch sensation Trans-Splicing Translation Initiation Translations Trypanosoma brucei brucei Work base chemotherapy in vivo insight interest mRNA Stability mutant novel pathogen preference prevent public health relevance research study sugar trafficking

项目摘要

DESCRIPTION (provided by applicant): The Order Kinetoplastida contains several pathogens of medical importance. These organisms employ an unusual mechanism of gene expression that requires the trans-splicing of every nuclear mRNA with the 39-nt spliced leader (SL) RNA. This RNA and mode of splicing is not found in humans and represents a unique target for chemotherapy. The SL RNA receives a m7G cap co-transcriptionally, followed by the addition of seven methylations on the sugar and/or base moieties of the first four nucleotides, constituting the unique 'cap 4'. Undermethylated substrate SL RNA is trafficked intracellularly prior to trans-splicing in Trypanosoma brucei and Leishmania tarentolae, introducing potential competitors into the cytosol for translation initiation factors. The basis of this application is a new addition to our hypothesis on the maturation of SL RNA involving a 'mask' protein that prevents eIF4F translation-initiation complex formation on the unspliced substrate. A unifying explanation for the undermethylation and loss of polysome formation for SL RNA mutants in L. tarentolae, which generally exhibit the cap 1 phenotype, is that they are recognized by the translational masking factor, but cannot be unmasked. The inability of these SL mutants to remove the translational mask is that 1) their 5' ends are not accessable for subsequent cap 2-4 methylations and 2) once they are trans-spliced, they are still translationally masked and either not exported from the nucleus or not loaded onto polysomes in the cytosol. The level of discrimination that exists among known cap-binding proteins has led us to search for this SL mask in the families of eIF4F factors. The hypothesis underlying these experiments is that immature SL RNA must be masked from the translation machinery so that the incomplete 5'-cap structure does not interfere with protein synthesis. The presence of six potential homologs of the cytosolic cap-binding protein eIF4E and six of associated translation initiation factor eIF4G in T. brucei provided candidates for examination; preliminary data indicates that TbeIF4E-3 fits the SL mask requirements. The specific aims of this application are: 1) To validate the cap-binding properties of TbeIF4E-3, a homolog of the Leishmania major cap 0-binding protein, and/or other candidates; and 2) To determine the TbeIF4E-3 RNA substrate(s) and any associated proteins. These studies will lead to a thorough understanding of kinetoplastid SL biogenesis and the interplay between RNA processing and translation. PUBLIC HEALTH RELEVANCE: The Order Kinetoplastida contains several pathogens of medical importance. These organisms employ an unusual mechanism of gene expression that requires the transsplicing of every nuclear mRNA with the 39-nt spliced leader (SL) RNA. This RNA and mode of splicing is not found in humans and represents a unique target for chemotherapy.

描述（由申请人提供）：命令动型层包含几种医学重要性的病原体。这些生物采用了基因表达的异常机制，需要用39-nt剪接的领导者（SL）RNA对每个核mRNA进行跨化。这种RNA和剪接模式在人类中找不到，代表了化学疗法的独特靶标。 SL RNA共同转录M7G CAP，然后在前四个核苷酸的糖和/或基部部分上添加7种甲基化，构成了独特的“ CAP 4”。甲基化的底物SL RNA在锥虫瘤和利什曼原虫塔伦氏菌进行移植之前被细胞内贩运，并将潜在的竞争者引入细胞质中，以进行翻译起始因子。该应用的基础是我们关于涉及涉及“掩模”蛋白的SL RNA成熟的假设的新补充，该蛋白可以防止未贴合的底物上的EIF4F翻译 - 引入复合物形成。对于通常表现出CAP 1表型的L. tarentolae中SL RNA突变体的甲基化不足和多甲基化形成的统一解释是，它们是通过翻译掩蔽因子认识到的，但不能透露。这些SL突变体无法去除翻译蒙版的能力是，1）它们的5'末端无法用于随后的2-4个甲基化的帽子，而2）一旦它们进行了移植，它们仍会被翻译掩盖，或者没有从细胞核中导出，或者没有从胞糖醇中载入多核细胞。在已知的帽结合蛋白中存在的歧视水平使我们在EIF4F因素的家族中寻找了这种SL面具。这些实验的基础假设是必须从翻译机制中掩盖未成熟的SL RNA，以使不完整的5'-cap结构不会干扰蛋白质的合成。胞质帽结合蛋白EIF4E的六个潜在同源物和六个相关的翻译起始因子EIF4G在T. brucei中提供了候选候选者；初步数据表明TBEIF4E-3适合SL面膜要求。本应用的具体目的是：1）验证Tbeif4e-3的盖结合特性，Tbeif4e-3，Leishmania主要CAP 0结合蛋白和/或其他候选者的同源物； 2）确定TBEIF4E-3 RNA底物和任何相关蛋白。这些研究将导致对动作塑料SL生物发生以及RNA处理和翻译之间的相互作用的透彻理解。公共卫生相关性：Kinetoplastida命令包含几种医学重要性的病原体。这些生物采用了基因表达的异常机制，需要用39-nt剪接的领导者（SL）RNA对每个核mRNA进行转化。这种RNA和剪接模式在人类中找不到，代表了化学疗法的独特靶标。