REPEAT-CONTAINING RNA BINDING PROTEINS OF TRYPANOSOMES

摘要 鞭毛动质体原生动物对公共卫生造成危害和经济负担 发展中世界的边缘化人口。非洲人和动物锥虫病的病原体， 布鲁氏锥虫引起一些研究最多，但最难控制和治愈的寄生性感染。 病媒控制失败，缺乏疫苗和有毒疗法，将责任推给了鉴定、分子 了解和验证新药靶点。从基础科学的角度来看，这些早期的 分支突起是线粒体RNA生物学、抗原变异、寄主-病原体的有价值的模型 相互作用、转录后调控和其他过程。上一个供资期间的主要调查结果 定位一个RNA结合五肽重复序列(PPR，35个氨基酸)蛋白质家族作为中央管道 控制线粒体基因的表达。螺旋-旋转-螺旋重复序列的独特能力组装成 超螺旋阵列识别扩展的和成分多样化的RNA序列使必要的 40多个PPR的作用。这些RNA“序列阅读器”通常包括锥虫特有的结构域。 将其他非特定的修饰酶承诺到它们的底物上，或阻止RNA降解和 分机。这个项目将阐明核编码的PPR控制线粒体的机制 通过将酶复合体和核糖体招募到转录本的方式在定义的转录水平上进行mRNA的生物发生和翻译 处理阶段。我们建立了三个优先领域，专注于PPR，使mRNA5‘端修饰成为可能， 3‘-腺苷/尿苷基化和翻译。建立在我们最近发现的基因特异性转录的基础上 在启动和核酸裂解前的mRNA处理中，我们确定了PPR因子作为“蛋白质帽”、“编辑” 传感器，“聚(A)结合蛋白，和潜在的翻译激活剂。我们的目标是剖析分子 整合这些功能并通过包含重复序列破译RNA识别原理的机制 蛋白质。通过阐明它们的RNA结合部位、结构、相互作用和遗传改变和结果 蛋白质工程，这个项目建立在我们在一个重要领域的势头上，并扩大了 病原体特定的关键过程。

ABSTRACT Flagellated kinetoplastid protozoans inflict public health hazards and economic burden predominately on marginalized populations in the developing world. The agents of African human and animal trypanosomiasis, Trypanosoma brucei sp., cause some of the most-studied, but least manageable and curable parasitic infections. Failing vector control, lack of vaccines and toxic therapeutics place the onus on identification, molecular understanding and validation of new drug targets. From the fundamental science perspective, these early branching Excavata represent valuable models of mitochondrial RNA biology, antigenic variation, host-pathogen interaction, post-transcriptional regulation, and other processes. Major findings in the previous funding period position a family of RNA binding pentatricopeptide repeat (PPR, 35 amino acids) proteins as the central conduit controlling mitochondrial gene expression. The unique ability of helix-turn-helix repeats to assemble into superhelical arrays recognizing extended and compositionally diverse RNA sequences enables the essential roles of more than 40 PPRs. Often including trypanosome-specific domains, these RNA “sequence readers” commit otherwise non-specific modification enzymes to their substrates, or block RNA degradation and extension. This project will elucidate the mechanisms by which nuclear-encoded PPRs govern mitochondrial mRNA biogenesis and translation via recruiting enzymatic complexes and ribosomes to transcripts at defined processing stages. We establish three priority areas focusing on PPRs that enable mRNA 5′ end modification, 3′ adenylation/uridylation, and translation. Building on our recent discoveries of gene-specific transcription initiation and exonucleolytic pre-mRNA processing, we identified PPR factors acting as “protein cap,” “editing sensor,” poly(A) binding protein, and potential translational activators. Our goal is to dissect the molecular machinery integrating these functions and to decipher principles of RNA recognition by repeat-containing proteins. By elucidating their RNA binding sites, structures, interactions, and outcomes of genetic alteration and protein engineering, this program builds on our momentum in a significant field and expands the knowledge of critical pathogen-specific processes.