The Role of Trypanosome Calcium Signaling in Parasite Virulence and Transmission

锥虫钙信号传导在寄生虫毒力和传播中的作用

• 批准号: 7546437
• 负责人: Brian T Emmer
• 金额: $ 2.77万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7546437
• 关键词: Address; Affinity; Animal Model; Antibodies; Antibody Formation; B-Lymphocytes; Back; Binding; Biochemical; Biological Assay; Biology; Blood; Blood Circulation; Calcium; Calcium Binding; Calcium Signaling; Calcium-Binding Proteins; Capsid Proteins; Cellular Assay; Chagas Disease; Characteristics; Class; Complex; Development; Disease; Engineering; Exhibits; Gene Targeting; Generations; Genetic; Genetic Techniques; Growth; Homologous Gene; Human; Humoral Immunities; Immune response; Immune system; In Vitro; Infection; Insect Vectors; Insecta; Investigation; Knockout Mice; Latin America; Life Cycle Stages; Lipids; Localized; Mammals; Mastigophora; Mediating; Membrane; Membrane Microdomains; Methods; Modeling; Modification; Molecular; Mus; Organism; Parasitemia; Parasites; Parasitic Diseases; Pathogenesis; Pathway interactions; Pharmaceutical Preparations; Phenotype; Population; Process; Proliferating; Proteins; Public Health; RNA Interference; Regulation; Research; Role; Signaling Protein; Staging; Stimulus; Structure; Study models; Testing; Trypanosoma; Trypanosoma brucei brucei; Trypanosoma cruzi; United States; Virulence; Work; cell motility; cell type; day; drug development; in vitro Assay; in vivo; mutant; programs; research study; response; restoration; surface coating; transmission process

DESCRIPTION (provided by applicant): The long-term objective of this application is to understand the molecular mechanisms by which Trypanosoma cruzi and Trypanosoma brucei are able to survive within the mammalian host and cause such profound disease. These flagellated protozoans have a complex life cycle, with programmed cellular differentiation regulating many aspects of parasite biology as the parasite transitions from the bloodstream of the mammalian host to the insect vector and back. This research focuses on the regulation and function of stage-specific parasite responses, with special emphasis on those enabling parasite survival and disease pathogenesis in the mammalian host. Specifically, this proposal investigates a major class of calcium signaling proteins in trypanosomes. The calflagins of T. brucei are studied in this proposal, because of genetic advancements in this organism enabling targeted gene inhibition by RNA interference. However, the conclusions derived are very likely to apply to the T. cruzi homologue as well (FCaBP), since they share >60% sequence identity, the same calcium-binding characteristics, lipid modifications, cellular localization, and lipid raft association. My preliminary work indicates that the calflagins (i) contribute to parasite virulence in murine models of infection, and (ii) are required for the differentiation from infectious forms of the parasite to transmissable forms. The first aim of this proposal seeks to identify the molecular pathway regulated by calflagin calcium-binding, through a combination of biochemical methods for purifying and characterizing interacting proteins and cellular assays for testing for deficits of calflagin mutants in hypothesized functions. The second aim seeks to determine the mechanism underlying the virulence and transmission deficit of calfalgin mutants. Because an interaction with host humoral immunity is suspected to underlie the virulence deficit of calfalgin mutants, the antibody response will be compared for calflagin RNAi and wild-type infections, and a restoration of normal virulence for calflagin mutants will be tested in B cell- deficient muMT knockout mice and in a setting of mixed co-infection with wild-type parasites. PUBLIC HEALTH RELEVANCE: These investigations have clear and direct implications to public health, as T. cruzi infects an estimated 8-11 million people in Latin America and another 100,000 in the United States. This research focuses on the ways in which these parasites are able to survive and cause disease in mammals. The proteins investigated in the proposed work, as well as interacting proteins identified by these experiments, are promising targets for drug development.

描述（由申请人提供）：本申请的长期目标是了解克氏锥虫和布氏锥虫能够在哺乳动物宿主体内存活并引起如此严重的疾病的分子机制。这些有鞭毛的原生动物具有复杂的生命周期，当寄生虫从哺乳动物宿主的血流转移到昆虫载体并返回时，程序化的细胞分化调节寄生虫生物学的许多方面。这项研究的重点是特定阶段寄生虫反应的调节和功能，特别强调那些使寄生虫在哺乳动物宿主中生存和疾病发病机制的反应。具体来说，该提案研究了锥虫中的一类主要钙信号蛋白。本提案对布氏锥虫的牛黄素进行了研究，因为该生物体的遗传进步使得通过 RNA 干扰能够实现靶向基因抑制。然而，得出的结论很可能也适用于 T. cruzi 同源物 (FCaBP)，因为它们具有 >60% 的序列同一性、相同的钙结合特征、脂质修饰、细胞定位和脂筏关联。我的初步工作表明，牛黄素（i）有助于小鼠感染模型中的寄生虫毒力，并且（ii）是将寄生虫的传染性形式区分为可传播形式所必需的。该提案的第一个目标是通过结合纯化和表征相互作用蛋白的生化方法以及测试钙旗素突变体在假设功能中的缺陷的细胞测定，来确定受钙旗素钙结合调节的分子途径。第二个目标旨在确定 calfalgin 突变体的毒力和传播缺陷的机制。因为与宿主体液免疫的相互作用被怀疑是calfalgin突变体毒力缺陷的基础，所以将比较calfalgin RNAi和野生型感染的抗体反应，并且将在B细胞缺陷的B细胞中测试calfalgin突变体正常毒力的恢复。 muMT 敲除小鼠以及与野生型寄生虫混合感染的环境。公共健康相关性：这些调查对公共健康具有明确而直接的影响，因为克氏锥虫感染了拉丁美洲约 8-1100 万人，以及美国另外 10 万人。这项研究的重点是这些寄生虫能够在哺乳动物中生存并引起疾病的方式。在拟议的工作中研究的蛋白质，以及通过这些实验鉴定的相互作用蛋白质，是药物开发的有希望的目标。