RNA CAPPING IN TRYPANOSOMES

锥虫中的 RNA 加帽

基本信息

批准号：
8260406
负责人：
CHRISTIAN TSCHUDI
金额：
$ 40.55万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-06-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8260406
关键词：
Adverse effects African African Trypanosomiasis Binding Biogenesis Biology Cattle Cell Death Cell physiology Central America Chagas Disease Code Communicable Diseases Complex Data Disease Domestic Pig Drug resistance Enzymes Family member Funding Genes Genetic Genome Goals Guanine Hand Health Human Human Bites Individual Infection Laboratories Leishmaniasis Life Life Cycle Stages Light Livestock Messenger RNA Metabolism Metals Methyltransferase Mini-Exons Modification N-terminal Nuclear Nuclear Protein Organism Parasite Control Parasites Pathway interactions Pharmaceutical Preparations Polyadenylation Polymerase Process Proteins Protozoa Public Health RNA RNA Caps RNA Polymerase II RNA Processing RNA triphosphatase Resistance Small Nuclear RNA South America Specificity Spliced Leader RNA Spliced Leader Sequences Staging Structure Therapeutic Toxic effect Trans-Splicing Transcript Treatment Cost Trypanosoma Trypanosoma brucei brucei Trypanosomatina Tsetse Flies Vaccines Work World Health Organization base burden of illness disability-adjusted life years guanylyltransferase in vivo interest mRNA Precursor nagana novel prevent protein protein interaction trait wasting

项目摘要

DESCRIPTION (provided by applicant): This application focuses on African trypanosomes of the Trypanosoma brucei subspecies, the causative agents of human African trypanosomiasis, a fatal disease unless treated, with a calculated disease burden of about two million Disability Adjusted Life Years. There are no vaccines and the few available therapeutic drugs have serious side effects and decreasing efficacy in light of the emergence of drug-resistant trypanosomes. Our interest in preventing and curing parasite infections is focused on understanding and ultimately exploiting genetic mechanisms that are essential for all stages of the parasite life cycle, but are either absent or fundamentally different in the human host. One possible therapeutic approach is suggested by the observation that trypanosomatids have strikingly unusual mRNA biosynthetic pathways. A key molecule in this process is the RNA polymerase (pol) II-transcribed spliced leader (SL) RNA, because its m7G-capped 5' terminal sequence is trans-spliced onto the 5' end of each mRNA. Furthermore, a subset of small nuclear RNAs involved in RNA processing, including U1, U2, U3 and U4, are also capped, but they are unusual in that the corresponding genes are transcribed by pol III. In the last funding period our work has led to the identification of two pathways that are essential for mRNA metabolism and viability in T. brucei. First, m7G capping of the SL RNA is carried out by a dedicated capping enzyme, TbCGM1, whose depletion leads to cell death. Second, the hypermodified cap 4 structure of mature SL RNA is specifically recognized by a nuclear cap-binding complex, which is unique in the eukaryotic kingdom and whose depletion is also incompatible with life. Furthermore, we have made the first step towards defining the cellular function of the previously identified capping enzyme TbCE1, namely m7G capping of some of the U-snRNAs; this is a distinctive trait not found in any other eukaryotic organism. Most interestingly, the N-terminal domain of TbCE1 harbors a metal-independent RNA triphosphatase activity with no structural or functional similarities to the previously characterized T. brucei RNA triphosphatase TbCet1 and both triphosphatases are fundamentally different from the triphosphatase present in the human host. The long- term goal of this proposal is to understand the parasite-specific biology and mechanisms of RNA capping in T. brucei and to identify essential factors, factor domains, or protein-protein interactions which might be exploited for parasite control. In the next funding period we plan to: 1. Carry out a functional analysis of the SL RNA m7G capping machinery and the novel metal-independent triphosphatase domain of TbCE1. 2. Identify determinants that enable selective capping of the pol II-transcribed SL RNA and a subset of pol III transcripts. 3. Further characterize the nuclear cap binding complex to elucidate the basis for its specificity for the cap 4 structure and define its in vivo function more precisely. PUBLIC HEALTH RELEVANCE: Parasitic protozoa are a major cause of global infectious diseases and thus, represent one of the most serious threats to public health. Among these are African trypanosomes, the causative agents of African trypanosomiasis or sleeping sickness in humans and a wasting and fatal disease (Nagana) in cattle, domestic pigs and other farm animals causing a profound effect on the economy of much of the continent. Unless treated, African sleeping sickness is always fatal; no vaccine has been approved and there is a very limited arsenal of drugs with generally severe shortcomings, such as high toxicity and emerging resistance.

描述（由申请人提供）：本申请的重点是锥虫亚种的非洲锥虫大体，即人类非洲锥虫病的致病药物，除非治疗，否则是一种致命的疾病，其计算出的疾病负担约为200万残疾人的寿命。没有疫苗，鉴于耐药性锥虫的出现，少数可用的治疗药物具有严重的副作用和降低功效。我们对预防和治愈寄生虫感染的兴趣集中在理解和最终利用对于寄生虫生命周期的所有阶段至关重要的遗传机制，但在人类宿主中没有或根本不同。观察到的一种可能的治疗方法表明，锥虫具有非常异常的mRNA生物合成途径。在此过程中的一个关键分子是RNA聚合酶（POL）II转录的剪接引导（SL）RNA，因为其M7G限制的5'末端序列被移至每个mRNA的5'端。此外，还限制了包括U1，U2，U3和U4在内的RNA处理中涉及的小核RNA的子集，但它们是不寻常的，因为相应的基因由POL III转录。在最后的资金期间，我们的工作导致了两种途径，这些途径对于mrna代谢和Brucei中的生存能力至关重要。首先，SL RNA的M7G上限是由专用封盖酶TBCGM1进行的，其耗竭导致细胞死亡。其次，成熟的SL RNA的高温盖4结构是由核帽结合复合物特别识别的，核帽结合络合物在真核界是独一无二的，其消耗也与生命不相容。此外，我们已经迈出了定义先前识别的封盖酶TBCE1的细胞功能的第一步，即某些U-SNRNA的M7G限额；这是在任何其他真核生物中都找不到的独特特征。最有趣的是，TBCE1的N末端结构域具有金属独立的RNA三磷酸酶活性，与先前表征的Brucei RNA Triphosphatase TBCET1无结构或功能相似性，并且两种三磷酸酶与存在于人类宿主中存在的TripHophatase的基本上都不同。该提案的长期目标是了解寄生虫特异性的生物学和Brucei中RNA限制的机制，并确定可能利用用于寄生虫控制的基本因素，因子域或蛋白质 - 蛋白质相互作用。在下一个资金期间，我们计划：1。对SL RNA M7G封盖机械和TBCE1的新型金属独立的三磷酸酶结构域进行功能分析。 2.确定确定性的决定因素，即可以选择POL II转录的SL RNA和POL III转录本的子集。 3。进一步表征核帽结合复合物，以阐明其对CAP 4结构的特异性的基础，并更精确地定义其体内功能。公共卫生相关性：寄生原生动物是全球传染病的主要原因，因此代表了对公共卫生最严重的威胁之一。其中包括非洲锥虫，非洲锥虫病的致病药物或人类昏昏欲睡的疾病，以及牛，家猪和其他农场动物的浪费和致命疾病（Nagana），对整个非洲大陆的经济产生了深远的影响。除非接受治疗，否则非洲睡眠始终是致命的。尚无疫苗已获得批准，并且具有通常严重缺陷的药物（例如高毒性和耐药性）的武器库非常有限。