Plasmodium cynomolgi as a model for P. vivax.

食蟹猴疟原虫作为间日疟原虫的模型。

• 批准号: 8290557
• 负责人: MARY R GALINSKI
• 金额: $ 17.6万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8290557
• 关键词: Address; Animal Model; Anopheles Genus; Applied Genetics; Area; Basic Science; Biochemical; Biochemistry; Biological; Biological Models; Biology; Blood; Cataloging; Catalogs; Caveolae; Cebidae; Cell membrane; Cells; Cellular biology; Cleaved cell; Color; Complex; Culicidae; Cytoplasm; Dependence; Development; Disease; Erythrocytes; Face; Future; Genetic; Genetic Transformation; Goals; Human; Immunology; In Vitro; Infection; Intervention; Investigation; Knowledge; Laboratories; Life Cycle Stages; Liver; Location; Macaca mulatta; Malaria; Membrane; Microscopy; Modality; Modeling; Morphology; Parasitemia; Parasites; Pathway interactions; Performance; Phylogenetic Analysis; Pilot Projects; Plasmodium cynomolgi; Plasmodium falciparum; Plasmodium vivax; Procedures; Proteins; Proteomics; Regulation; Research; Resources; Reticulocytes; Rodent; Sporozoites; Staging; Surface; System; Technology; Tertiary Protein Structure; Therapeutic; Time; Transfection; Vacuole; Vesicle; biological research; experience; feeding; in vivo; in vivo Model; interest; model development; novel; pathogen; programs; protein transport; research study; success; tool; trafficking; vaccine development

DESCRIPTION (provided by applicant): This R21 proposal focuses on developing in vivo and in vitro Plasmodium cynomolgi model systems to advance cell biological and biochemical research on P. vivax, a major widespread human malaria pathogen. In particular, the goal is to enable the routine application of transfection technologies that will address hypothesis-driven questions regarding the unique biology held in common between P. vivax and P. cynomolgi, which is not found in the other major human malaria, P. falciparum or any of the popular rodent malaria models. This biology includes, among many other differences, the development and reactivation of dormant liver-stage forms known as hypnozoites, and the synthesis and functioning of numerous caveolae vesicle complexes (CVCs) and other membrane structures in infected red blood cells (iRBCs). While P. vivax cannot be cultured continuously in vitro, to facilitate such experimentation, P. cynomolgi blood-stage parasites can be easily manipulated genetically in vivo and ex vivo from rhesus monkey infections where large quantities of parasites can be obtained as well as be adequately propagated and experimentally manipulated by in vitro culture for several days to weeks. The phylogenetic close kinship and nearly identical basic morphology and biology shared by these two species strongly support the proposed utility of P. cynomolgi model systems. No rodent malaria model or P. falciparum culture system offers any biology that mimics the unique biology of these species, and thus they cannot serve as model systems to forward this research. In Aim 1 we will focus on attaining transformed P. cynomolgi parasites that constitutively throughout the life-cycle or at specific developmental time points express single or dual-color fluorescent tags, as the first step for future studies aiming to identify, purify and investigate specific life cycle stages, with a primary interest at this time on the development and activation of the elusive hypnozoite stage. In Aim 2, transformed parasites will be developed to study the trafficking machinery, pathways and unique membrane structures produced by P. vivax and P. cynomolgi blood-stage parasites, which include the CVCs, extensive networks of cleft membranes and novel proteins catalogued by proteomics, immunochemical studies and microscopy. One specific question to be addressed is how and when the PHIST81-95 protein traffics from the parasite beyond the parasitophorous vacuole membrane to its RBC cytosolic location on the cytoplasmic face of the CVCs. For this, the Conditional Aggregation Domain (CAD) protein regulation system will be applied. Critically, P. cynomolgi parasites provide a superior model for the proposed studies, and development of this model will enable research on initial hypothesis-driven questions posed here and ongoing research to understand and define biological and biochemical targets of intervention for P. vivax.

描述（由申请人提供）：该R21提案侧重于开发体内和体外食蟹猴疟原虫模型系统，以推进对间日疟原虫（一种主要的广泛传播的人类疟疾病原体）的细胞生物学和生物化学研究。具体而言，目标是使转染技术的常规应用成为可能，这将解决关于间日疟原虫和食蟹猴疟原虫之间共同拥有的独特生物学的假设驱动的问题，这在其他主要的人类疟疾、恶性疟原虫或任何流行的啮齿动物疟疾模型中都没有发现。这种生物学包括，在许多其他差异中，休眠的肝脏阶段形式（称为催眠虫）的发育和重新激活，以及许多小窝囊泡复合物（CVC）和感染的红细胞（iRBC）中其他膜结构的合成和功能。虽然间日疟原虫不能在体外连续培养，但为了促进这样的实验，食蟹猴疟原虫血液阶段寄生虫可以容易地在体内和离体从恒河猴感染中进行遗传操作，其中可以获得大量寄生虫，并且通过体外培养数天至数周来充分繁殖和实验操作。这两个物种的系统发育密切的亲缘关系和几乎相同的基本形态和生物学，强烈支持建议的实用性P. cynomolgi模式系统。没有啮齿动物疟疾模型或恶性疟原虫培养系统提供任何模拟这些物种的独特生物学的生物学，因此它们不能作为推进这项研究的模型系统。在目标1中，我们将专注于获得转化的食蟹猴疟原虫寄生虫，这些寄生虫在整个生命周期中或在特定的发育时间点表达单色或双色荧光标签，作为未来研究的第一步，旨在鉴定，纯化和研究特定的生命周期阶段，此时主要关注难以捉摸的催眠虫阶段的发育和激活。在目标2中，将开发转化的寄生虫以研究由间日疟原虫和食蟹猴疟原虫血液阶段寄生虫产生的运输机制、途径和独特的膜结构，其中包括CVC、广泛的裂膜网络和通过蛋白质组学、免疫化学研究和显微镜编目的新蛋白质。要解决的一个具体问题是PHIST 81 -95蛋白如何以及何时从寄生虫运输到寄生虫空泡膜之外的CVC细胞质表面上的RBC胞质位置。为此，将应用条件聚集结构域（CAD）蛋白质调节系统。重要的是，食蟹猴疟原虫寄生虫为所提出的研究提供了一个上级模型，该模型的开发将使对本文提出的初始假设驱动问题的研究和正在进行的研究能够理解和定义间日疟原虫干预的生物学和生化靶标。