Functional analysis of molecular components of the habitat of Leishmania spp. - the protozoan parasites causing human leishmaniases

利什曼原虫是医学上重要的细胞内变形动物寄生虫。使用蛋白质组学方法，我们之前全面分析了寄生虫对巨噬细胞内细胞内栖息地的适应。在这里，我们建议通过分析宿主细胞来源的膜和可溶性蛋白质来扩展和补充这种方法。具体地说，我们建议在含有利什曼原虫和含有乳胶珠的吞噬体之间进行宿主吞噬体多蛋白复合物的比较硅分析，以确定功能上的相关差异。此外，我们将探索一个已经确定的差异的相关性，并验证以下假设：1)利什曼- pv的胆固醇/脂质积累，2)宿主细胞胆固醇合成途径的上调，3)膜筏依赖信号级联的抑制，4)寄生虫脂肪酸分解代谢的上调是机械联系和相互依赖的。最后，我们同时希望建立基于胚胎干细胞和转基因小鼠来源的原代吞噬细胞的实验平台，这将允许利用反向遗传学或RNAi的力量来分析不同细胞背景下的吞噬体蛋白功能。

Leishmania spp. are medically important intracellular proteozoan parasites. Using a proteomics approach we have previously comprehensively analysed parasite adaptation to its intracellular habitat in macrophages. Here we propose to extend and complement this approach by analysing the host cell-derived membrane and soluble proteins that contribute to this habitat. Specifically we propose to perform a comparative in silico analysis of host phagosomal multiprotein complexes between Leishmania- and latex bead-containing phagosomes to define functionally relevant differences. . Furthermore, we will pursue the relevance of one already identified difference and test the hypothesis that i) Cholesterol/lipid accumulation in Leishmania-PVs, ii) up-regulation of host cell cholesterol synthesis pathways, iii) inhibition of membrane raft dependent signalling cascades and iv) up-regulation of fatty acid catabolism in parasites are mechanistically linked and interdependent. Finally, we simultaneously want to establish experimental platforms based on ES cell- and transgenic mouse-derived primary phagocytic cells that will allow to harness the power of reverse genetics or RNAi to analyse phagosomal protein function in different cell contexts.