Function and Trafficking of Flagellar Membrane Proteins in Leishmania mexicana

利什曼原虫和其他动体寄生虫会导致毁灭性的疾病，折磨着数百万人。全 动体寄生虫在生命周期的所有阶段都含有鞭状细胞器。研究结束 几十年来，鞭毛在这些寄生虫的生物学中扮演着至关重要的不同角色， 包括运动，附着在昆虫媒介的肠道上，感知环境，调节交配， 与哺乳动物宿主细胞内膜相互作用，逃避宿主免疫反应等 这样的鞭毛对于寄生虫在昆虫媒介中的定植和传播以及对 脊椎动物宿主内的生存和毒力。而鞭毛的内部组件，如 轴丝，已被更详细地研究，鞭毛膜(FM)的表面知之甚少， 尽管FM介导了上述大多数生物学功能。为了更好地了解FM和 它在寄生中的作用，这项提议将调查：目的1)特定的FM蛋白如何选择性地向 表面膜的离散成分；目的2)几个选定的FM的生物学功能是什么 蛋白质。该项目建立在对Kharon蛋白的初步研究基础上，Kharon蛋白介导了FM的贩运 鞭毛葡萄糖转运体GT1，一种对细胞内疾病的生存至关重要的运输机器- 引起无鞭毛体阶段的。Kharon转运GT1到鞭毛内转运的分子机制 将对运送到鞭毛中的颗粒进行测试。由于大多数FM蛋白不依赖于Kharon 对于FM交易，与这些FM蛋白质相互作用的其他FM交易机器将通过 串联亲和纯化/质谱法。然后将对这些新型FM贩运机进行调查， 通过基因敲除方法，在昆虫载体和哺乳动物感染中的关键功能 生命周期的各个阶段。三种选定的具有已知或可能生化功能的FM蛋白将被 通过基因敲除，研究昆虫媒介和哺乳动物宿主阶段的功能。这些研究 将揭示FM的特定分子成分如何在两种情况下调节寄生虫的关键功能 昆虫媒介和哺乳动物宿主。

Leishmania and other kinetoplastid parasites cause devastating diseases that afflict millions of people. All kinetoplastid parasites contain a whip-like organelle, the flagellum, in all life cycle stages. Studies over decades have underscored the vital and diverse roles that flagella play in the biology of these parasites, including motility, attachment to the gut of insect vectors, sensing the environment, mediating mating, interacting with internal membranes of mammalian host cells, evading the host immune response, etc. As such, flagella are essential for both parasite colonization of and transmission through the insect vector and for survival and virulence inside the vertebrate host. While internal components of the flagellum, such as the axoneme, have been studied in greater detail, the surface of the flagellar membrane (FM) is poorly understood, despite the fact that the FM mediates most of the above biological functions. To better understand the FM and its role in parasitism, this proposal will investigate: Aim 1) how specific FM proteins traffic selectively to this discrete component of the surface membrane; Aim 2) what the biological functions are for several selected FM proteins. The project is founded on initial studies on the KHARON protein, that mediates FM trafficking of a flagellar glucose transporter GT1, a trafficking machine that is essential for viability of intracellular disease- causing amastigote stages. A molecular mechanism for KHARON in delivering GT1 to Intraflagellar Transport particles for trafficking into the flagellum will be tested. Since most FM proteins do not depend upon KHARON for FM trafficking, other FM trafficking machines that interact with those FM proteins will be identified by tandem affinity purification/mass spectrometry. These novel FM trafficking machines will then be investigated, by gene knockout approaches, for critical functions in both the insect vector and the mammalian infective stages of the life cycle. Three selected FM proteins with known or likely biochemical functions will then be investigated, by gene knockout, for functions in the insect vector and mammalian host stages. These studies will reveal how specific molecular components of the FM mediate critical functions for the parasite in both the insect vector and the mammalian host.