Control of stage differentiation of Trypanosoma brucei by the host environment: a large scale RNAi screen of the kinome and phosphoproteomic analysis

锥虫穿梭于脊椎动物和吸血昆虫之间，非洲热带的采采蝇。在寄生生活史中，迅速适应宿主环境的突然变化是生存的关键。宿主环境提供的信号触发寄生虫基因表达的协调变化，导致分化为适应的发育阶段。我们以前的工作表明，温度变化（冷休克），柠檬酸或cisaconitate在培养基中，和细胞周期停滞的巧合是重要的启动分化的苍蝇中肠阶段。我们建议确定一个尚未探索的途径，传输主机线索温度和柠檬酸盐的分子组成部分。两种互补的方法将继续进行：（1）筛选一组生物信息学鉴定的207个候选基因，包括T。在诱导分化的培养模型中使用RNAi技术对布鲁氏菌基因组进行了研究;（2）在2D蛋白电泳中调查了由宿主线索诱导的蛋白磷酸化变化，并通过磷酸化蛋白质组学鉴定了其他途径组分;（3）表征了已经发现的冷诱导蛋白激酶。我们期待不寻常的和新的信号蛋白，锥虫的早期系统发育分支。因此，深入了解信号和interorganisms控制机制的演变，并发现潜在的药物靶点，目前不充分的治疗重要的热带疾病可能会导致这个项目。

Trypanosomes shuttle between vertebrates and a blood feeding insect, the tsetse in tropical Africa. In the parasitic life cycle, rapid adaptation to sudden changes of the host environments are essential for survival. Signals provided by the host environment trigger coordinate changes in parasite gene expression that result in differentiation to adapted developmental stages. Our previous work shows that coincidence of temperature change (cold shock), citrate or cisaconitate in the medium, and cell cycle arrest is important to initiate differentiation to the fly midgut stage. We propose to identify molecular components of a yet unexplored pathway that transmits the host cues temperature and citrate. Two complementary approaches will be pursued: (1) screening of a bioinformatically identified set of 207 candidate genes, including all protein kinases in the T. brucei genome, using RNAi in a culture model of induced differentiation; (2) survey of protein phosphorylation changes induced by the host cues in 2D protein electrophoresis, and identification of additional pathway components by phosphoproteomics; (3) characterization of an already discovered cold-induced protein kinase. We expect unusual and novel signaling proteins, given the early phylogenetic branching of trypanosomes. Consequently, insights into the evolution of signaling and interorganismic control mechanisms, and discovery of potential drug targets for presently insufficient therapy of important tropical diseases may result from this project.