Collaborative Research: RNA Processing in Trypanosome Mitochondria

该项目将研究古代单细胞寄生虫锥虫基因表达的独特机制。在大多数生物体中，RNA是由DNA复制而来，在不改变密码的情况下指导蛋白质合成。然而，在锥虫体线粒体（细胞的“发电站”）中，RNA通过添加或去除特定核苷酸而被广泛编辑。在其复杂的生命周期中，锥虫在两个非常不同的宿主之间交替：人类（和其他动物）和昆虫，后者是传播媒介。这种寄生虫在人类和昆虫中所面临的截然不同的环境需要快速和大规模的代谢和生理变化，包括RNA编辑。在寄生虫发育过程中，RNA编辑机制是如何被精确调控的关键问题仍然没有答案，这也是本研究的重点。该项目将通过为博士后、研究生、本科生和高中生提供跨学科培训机会，重点关注传统上在STEM学科中代表性不足的群体，从而产生广泛的教育影响。本项目重点研究布鲁氏锥虫mRNA编辑调控以及调控编辑体中编辑保真度的REH2C复合体的关键作用。这项研究可以为一个几十年来一直是个谜的问题提供新的线索：在不同的布鲁氏虾生命周期中，mRNA的编辑是如何受到不同的控制的？目的1将测试“部分编辑控制”（PEC）模型，利用部分编辑来差异抑制两种发育形式的布氏体mRNA成熟。最近开发的RNA-seq/生物信息学分析编辑rna的管道将用于此目的。目的2将探讨编辑中底物特异性的来源，特别是定义一种罕见的DEAH-box解旋酶辅助因子及其个体锌指在该机制中的作用。目的3将通过使用遗传、生化（接近标记）和化学（交联质谱）方法阐明编辑体的一般组织，研究编辑成分和REH2C复合体重塑的动态相互作用。总的来说，这些研究将揭示寄生虫两个生命周期阶段编辑调控的新方面，包括编辑保真度、底物特异性决定因素和编辑体相互作用。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This project will study a distinctive mechanism of gene expression in ancient single-celled parasites called trypanosomes. In most organisms, RNA is copied from DNA and directs protein synthesis without changes in the code. However, in trypanosomal mitochondria (the ‘powerhouse’ of cells), RNA is edited extensively by addition or removal of specific nucleotides. In their complex life cycle, trypanosomes alternate between two very different hosts: humans (and other animals) and insects, which are the vectors of tranmission. The dramatically different environments that the parasites face in humans and insects demand rapid and large-scale metabolic and physiological changes, including in RNA editing. The crucial question of how the RNA editing mechanism is precisely regulated during parasite development remains unanswered and is the focus of this research. The project will have broad educational impact by providing interdisciplinary training opportunities for postdoctoral, graduate, undergraduate, and high school students, with a focus on groups traditionally underrepresented in the STEM disciplines.This project focuses on Trypanosoma brucei mRNA editing regulation and the key role of the REH2C complex that controls editing fidelity in editosomes. The research can shed new light on a question that has remained a mystery for decades: how is mRNA editing differentially controlled across the T. brucei life-cycle? Objective 1 will test the “partial-editing control” (PEC) model of exploiting partial editing to differentially inhibit mRNA maturation in two developmental forms of T. brucei. A recently developed pipeline for RNA-seq/bioinformatic analysis of edited RNAs will be employed for this purpose. Objective 2 will address the source of substrate specificity in editing, specifically to define the role of a rare DEAH-box helicase cofactor and its individual zinc fingers in the mechanism. Objective 3 will examine dynamic interactions of editing components and remodeling by the REH2C complex by elucidating the general organization of editosomes using genetic, biochemical (proximity labeling), and chemical (crosslinking-mass spectrometry) approaches. Overall, these studies will reveal new aspects of editing regulation in two life-cycle stages of the parasite, including editing fidelity, substrate specificity determinants, and editosome interactions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.