Control of VSG pre-mRNA processing in infectious Trypanosoma brucei

项目摘要 这一应用重点是原生动物寄生虫布鲁氏锥虫，这种寄生虫在 哺乳动物通过采采蝇寄主，由于有效的免疫逃避，会导致慢性和致命性感染， 特别是非洲人的昏睡病和牛的纳加纳病。而人类非洲锥虫病 最近一直在下降，牲畜感染仍然很普遍，并对经济产生了深远的影响 撒哈拉以南非洲的发展。没有疫苗和治疗药物有严重的副作用和 药效下降。因此，迫切需要进行研究，以更好地了解这些疾病的生物学 病原体和它们用来在宿主体内生存的机制。布氏锥虫经历了一个复杂的生活史 在哺乳动物宿主和食血采采蝇媒介之间。为了应对不断变化的环境， 寄生虫通过进化以确保生存和成功的不同的生命周期形式进行过渡 传输到下一台主机。例如，当居住在哺乳动物宿主时，布氏毛滴虫表达一种 变异型表面糖蛋白(VSG)外壳，这是抗原变异的范例。尽管有 在基因组中数百个VSG基因中，血流形式的VSG表达被限制在大约15个基因中的1个 专门的端粒血流表达位点(BES)。在昆虫媒介变环虫的唾液腺中 锥体被VSG的一个特定的小子集所覆盖，即亚环VSG(MVSG)，它 使其能够传播到脊椎动物宿主。MVSG表达由未知机制触发，并且在 在没有采采树组织的情况下，可以通过诱导RNA结合蛋白6的表达来实现体外培养 (RBP6)。这项应用的一个主要目标是检查锥虫是如何接收指令开始的 合成mVSG外壳，以及当每个细胞开始表达单个mVSG时。我们将使用单细胞 RNA-Seq和长期活细胞成像来回答机械问题。另外，我们的新发现是 编码外显子剪接增强子的mVSG基因揭示了VSG调控的一个全新方面 这可能解释了成熟mRNA为什么受到严格控制的极高产量。

Project Summary This application focuses on the protozoan parasite Trypanosoma brucei, which is transmitted among mammalian hosts by tsetse-flies and, due to effective immune evasion, causes chronic and lethal infections, specifically African sleeping-sickness in humans and nagana in cattle. Whereas human African trypanosomiasis has been declining recently, livestock infections remain prevalent and have a profound effect on economic development 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 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. To cope with the changing environment, the parasite transitions through distinct life cycle forms that have evolved to assure survival and successful transmission to the next host. For instance, when residing in the mammalian host, T. brucei expresses a variant surface glycoprotein (VSG) coat, which is the paradigm for antigenic variation. Although there are hundreds of VSG genes in the genome, bloodstream-form VSG expression is restricted to 1 of about 15 specialized telomeric bloodstream expression sites (BES). In the salivary glands of the insect vector metacyclic trypanosomes are covered by a specific and small subset of VSGs, the metacyclic VSGs (mVSGs), which enable transmission to a vertebrate host. mVSG expression is triggered by an unknown mechanism and in vitro can be achieved in the absence of tsetse tissues by induced expression of the RNA-binding protein 6 (RBP6). One major goal of this application will be to examine how trypanosomes receive instructions to begin synthesizing the mVSG coat, and when each cell begins expressing a single mVSG. We will use single-cell RNA-Seq and long-term live-cell imaging to answer mechanistic questions. In addition, our novel finding that mVSG genes encode an exonic splicing enhancer exposed a completely new aspect of VSG regulation and may explain the tightly-controlled extremely high output of mature mRNAs.