Biosynthesis and Function of Base J

J碱基的生物合成和功能

• 批准号: 7032606
• 负责人: ROBERT S SABATINI
• 金额: $ 35.78万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-12-15 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7032606
• 关键词: DNA damage; RNA interference; Trypanosoma brucei; chromatin; developmental genetics; gene induction /repression; gene targeting; glycoprotein structure; glycosylation; host organism interaction; immune tolerance /unresponsiveness; laboratory mouse; molecular site; nucleic acid biosynthesis; nucleic acid repetitive sequence; nucleic acid structure; nucleobase; nucleosomes; parasite infection mechanism; protein protein interaction; protein structure function; protozoal genetics; surface antigens; telomere; thymine; transfection /expression vector

DESCRIPTION (provided by applicant): African trypanosomes (Trypanosoma brucei) are protozoan parasites that cause a fatal disease known as African sleeping sickness in humans and related diseases in animals. These diseases have devastating health and economic consequences. Pathogenesis of T. brucei is critically dependent on the phenomena of antigenic variation. During infection of the mammalian host the majority of the trypanosome population is destroyed by the immune response to the Variant Surface Glycoprotein (VSG) coat. In the process of antigenic variation, individual trypanosomes are able to switch the VSG gene that is expressed, from a repertoire of several hundred different VSG genes, and evade immune destruction. To be expressed, a VSG gene has to be located within a telomeric expression site (ES). However, there are approximately 20 ESs and only one is active at a time. The mechanism of selective expression/repression and switching of silent copies of VSG is not entirely understood. Previous analysis indicates that regulation of a given VSG ES correlates with the absence or presence of a uniquely modified DMA base, represented by the glycosylation of thymine residues and called base J. The association of J with transcriptional repression of telomeric VSG genes suggests its role in regulation of antigenic variation. In this proposal we aim to examine the molecular mechanisms underlying J activity by defining the protein/enzymatic activities associated with J and the consequences of J incorporation within the chromosomes of bloodstream form trypanosomes. Towards this end we have isolated a protein, JBP2, that regulates J-biosynthesis, determining both the developmental stage-specific and telomeric site-specific synthesis of the modified base. Preliminary analysis indicates that JBP2 is an essential gene suggesting that base J is an essential modification of the bloodstream chromosome. The broad, long-term objectives of this proposal are to use the biochemical and genetic analyses of JBP2 function as tools to advance ourunderstanding of the mechanism and biology of J-function in T. brucei. These analyses will allow us to directly address the role of telomeric DMA glycosylation on the regulation of antigenic variation. The potentially significant role of J to the survival of the parasite, and the absence of base J in their mammalian hosts, indicates attractive targets for the design of inhibitors with broad specificity and low toxicity. Enzymes and co-factors involved in J-biosynthesis would represent such targets. These studies may prove useful in identifying novel approaches to prevention, treatment and diagnosis of the debilitating and deadly diseases caused by these parasites.

描述（由申请人提供）：非洲锥虫（Brucei锥虫）是原生动物的寄生虫，引起致命疾病，称为人类中的非洲睡眠疾病，并在动物中引起相关疾病。这些疾病具有毁灭性的健康和经济后果。布鲁氏菌的发病机理至关依赖于抗原变异现象。在哺乳动物宿主的感染期间，大多数锥虫种群因对变异表面糖蛋白（VSG）涂层的免疫反应而破坏。在抗原变异的过程中，单个锥虫能够从数百种不同VSG基因的曲目中切换所表达的VSG基因，并逃避免疫破坏。为了表达，VSG基因必须位于端粒表达位点（ES）中。但是，大约有20个ESS，一次只有一个活跃。选择性表达/抑制和VSG无声拷贝的切换的机制尚不完全了解。先前的分析表明，给定VSG ES的调节 与不存在或存在唯一修饰的DMA碱基相关，该基碱由胸腺嘧啶残基的糖基化表示，并称为基础J。J的缔合与端粒VSG基因的转录抑制作用表明其在调节抗原变异的调节中的作用。在此提案中，我们旨在通过定义与J相关的蛋白质/酶促活性以及J掺入在血液形式形式的锥虫中的蛋白质/酶促活性来检验的分子机制。为此，我们分离了一种调节J-生物合成的蛋白质JBP2，从而确定了修饰碱的发育阶段特异性和端粒特异性合成。初步分析表明JBP2是一个必不可少的基因，表明碱基J是血液染色体的必要修饰。该提案的广泛，长期目标是将JBP2的生化和遗传分析用作提高我们对T. Brucei J功能机制和生物学的理解的工具。这些分析将使我们能够直接解决端粒DMA糖基化对抗原变异调节的作用。 J对寄生虫生存的潜在重要作用以及在其哺乳动物宿主中缺乏碱基J的潜在作用，这表明具有广泛特异性和低毒性抑制剂的设计有吸引力的靶标。参与J-Biosynsis的酶和共同因素将代表此类靶标。这些研究可能被证明可用于识别这些寄生虫引起的衰弱和致命疾病的新型预防，治疗和诊断的新方法。