Mechanism of Infectivity Acquisition in African Trypanosomes

项目概要 揭示人类病原体的生物学是理解病原体机制的基础 发病机制并鉴定宿主生存必需的基因。该应用程序重点关注 原生动物寄生虫布氏锥虫，在人类和动物中引起毁灭性的疾病 撒哈拉以南非洲。没有疫苗，治疗药物副作用严重 功效降低。因此，迫切需要进行研究以更好地了解这些物质的生物学特性。 人类病原体及其在宿主体内生存的机制。 T. brucei 经历了 哺乳动物宿主和吸血采采蝇媒介之间复杂的生命周期，其中 其他涉及细胞形态、表面涂层成分、新陈代谢、信号传导途径的变化 和基因表达。因此，这些寄生虫已经进化出适应能力以使其生存 在采采蝇的肠道和唾液腺以及哺乳动物的血液中 主机。通过在非传染性锥虫中过表达单个 RNA 结合蛋白 (RBP6)，我们 在体外概括了导致昆虫媒介获得感染性的事件，包括 后环变异表面糖蛋白（mVSG）的表达。拟议工作的总体目标 这里的目的是确定 RBP6 激活感染性发育进程的机制 元环化合物并表征调节发育的RNA结合蛋白网络 导致上鞭毛体和传染性后循环体的程序。为了实现这些目标，我们将在此基础上 我们在上一个资助期间的主要发现，即确定 RBP6 的初始目标 功能、后环化合物分子特征的描述以及大环化合物的识别 与导致感染性后循环寄生虫的发育程序相关的分子纲要。 我们将使用创新方法，包括通过深度突变扫描筛选 RBP6 功能 通过结合元循环选择和高通量 DNA 测序并分析基因表达 RNA-Seq 的单细胞水平。总的来说，我们的研究计划提供了独特的机会 阐明从非感染性前环化合物到感染性后环化合物的发展程序， T. brucei 生命周期中的关键过程。

Project Summary Unraveling the biology of human pathogens is fundamental toward understanding mechanisms of pathogenesis and identifying genes essential for survival in the host. This application focuses on the protozoan parasite Trypanosoma brucei, which causes devastating diseases in humans and animals in sub-Saharan Africa. There are no vaccines, and therapeutic drugs have serious side effects and decreasing efficacy. Thus, there is a pressing need for research to better understand the biology of these human pathogens and the mechanisms they use to survive within their hosts. T. brucei undergoes a complex life cycle between the mammalian host and the blood-feeding tsetse fly vector, which among others involves changes in cell morphology, surface coat composition, metabolism, signaling pathways and gene expression. Consequently, these parasites have evolved adaptations to allow for their survival in both the gut and salivary glands of the tsetse fly, as well as in the bloodstream of their mammalian host. By overexpressing a single RNA-binding protein (RBP6) in non-infectious trypanosomes, we recapitulated in vitro the events leading to acquisition of infectivity in the insect vector, including the expression of metacyclic variant surface glycoproteins (mVSGs). The overall goal of the proposed work here is to identify the mechanism by which RBP6 activates developmental progression to infectious metacyclics and to characterize the RNA binding protein network that regulates the developmental program leading to epimastigotes and infectious metacyclics. To accomplish these goals, we will build on our major findings in the previous funding period, namely the identification of initial targets of RBP6 function, the depiction of the molecular characteristics of metacyclics, and the identification of a large compendium of molecules linked to the developmental program leading to infective metacyclic parasites. We will use innovative approaches involving a screen for RBP6 function with deep mutational scanning by combining metacyclic selection and high-throughput DNA sequencing and analyze gene expression at the single cell level with RNA-Seq. Taken together our research plan provides unique opportunities to illuminate the developmental program leading from non-infective procyclics to infectious metacyclics, a crucial process in the T. brucei life cycle.