Proteomics of the Plasmodium Ookinete

项目描述（申请人提供）：本项目将应用蛋白质组学的先进和新兴技术，发现阻断疟疾传播的新分子靶点。虽然主要是探索性的，这一建议也是基于两个假设：1）疟原虫合子（蚊子中肠配子融合/受精的产物）表达新的、可能丰度低的、但迄今尚未鉴定的蛋白质，这些蛋白质可能是阻断疟疾传播的免疫学靶点;和2）疟原虫动卵细胞（受精卵之后的能动发育阶段，并积极侵入中肠）表达新的分泌和细胞表面-相关的分子，特别是参与寄生虫入侵蚊子中肠上皮细胞和阻断疟疾传播的潜在目标。有两个具体目标：1）利用多维蛋白质鉴定技术（MUDPIT）、定量同位素编码亲和标签质谱和定量2D凝胶电泳等互补技术，对鸡疟原虫受精卵和动合子的蛋白质组进行了描述，并利用生物信息学工具对现存疟原虫基因组数据库中受精卵和动合子阶段的基因进行了比较鉴定，特别是在致死性人恶性疟原虫中;和2）通过在传播阻断免疫的动物模型中测试选择数量的新鉴定的分子，用新的DNA疫苗技术和传统的重组蛋白表达/疫苗接种来开始受精卵/动合子表达的蛋白的验证。数据将被放入PlasmoDB公共数据库。本申请直接涉及父R 01授权AI 45999，“阻断疟疾传播的分子靶标”，因为本申请的重点是使用蛋白质组学来发现和验证阻断疟疾传播的新分子靶标。该提案也非常适合由R 01赠款支持的强大的现存基础设施，以测试新发现的寄生虫分子作为阻断疟疾传播的目标。该应用与R21机制的探索/开发性质一致，因为它提出应用新的和新兴的技术来发现未知但潜在重要和有趣的疟原虫分子。

DESCRIPTION (provided by applicant): This project will apply advanced and emerging technologies in proteomics to the discovery of novel molecular targets of blocking malaria transmission. While primarily exploratory, this proposal is also based on two hypotheses: 1) Plasmodium zygotes (the products of gamete fusion/fertilization in the mosquito midgut) express novel, perhaps low abundance, but hitherto unidentified proteins that may be immunological targets of blocking malaria transmission; and 2) Plasmodium ookinetes (the motile developmental stage that succeeds the zygote and actively invades the midgut) express novel secreted and cell surface-associated molecules that are specifically involved in parasite invasion of the mosquito midgut epithelium and are potential targets of blocking malaria transmission. There are two Specific Aims: 1) To delineate the proteome of Plasmodium gallinaceum zygotes and ookinetes using complementary techniques of multidimensional protein identification technology (MUDPIT), quantitative isotope-coded affinity tag mass spectrometry, and quantitative 2D gel electrophoresis, and use bioinformatics tools to comparatively identify zygote and ookinete stage genes in the extant Plasmodium genome databases, particular in the lethal human malaria parasite P. falciparum; and 2) To initiate the validation of zygote/ookinete-expressed proteins by testing a select number of newly identified molecules in an animal model of transmission-blocking immunity, both with a new DNA vaccine technology and traditional recombinant protein expression/vaccination. Data will be placed into the PlasmoDB public database. This application directly relates to the parent R01 grant AI45999, "Molecular Targets of Blocking Malaria Transmission," since the focus of the present application is on using proteomics to discover and validate new molecular targets of blocking malaria transmission. This proposal also fits well into the robust extant infrastructure supported by the parent R01 grant to test newly identified parasite molecules as targets of blocking malaria transmission. This application is consistent with the exploratory/development nature of the R21 mechanism since it proposes to apply new and emerging technologies to discovering unknown but potentially important and interesting Plasmodium molecules.