Characterization of calcium signaling proteins in Toxoplasma gondii

弓形虫钙信号蛋白的表征

• 批准号: 8352190
• 负责人: Gustavo A Arrizabalaga
• 金额: $ 7.8万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8352190
• 关键词: Affect; Antibiotic A23187; Calcium; Calcium Signaling; Cells; Cessation of life; Complement; Congenital Abnormality; Data Set; Defect; Development; Disease; Drug Delivery Systems; Event; Fetus; Funding Mechanisms; Genes; Genetic; Genome; Goals; Human; Immunocompromised Host; Individual; Invaded; Ionophores; Laboratories; Lead; Learning; Methods; Missense Mutation; Movement; Mutate; Mutation; Names; Nucleotides; Opportunistic Infections; Orphan; Parasites; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Population; Process; Proteins; Regulation; Signal Pathway; Signal Transduction; Signaling Protein; Sodium-Hydrogen Antiporter; Techniques; Therapeutic; Toxoplasma gondii; Toxoplasmosis; Work; calcium-dependent protein kinase; cell motility; combat; computerized data processing; exhaustion; extracellular; gene function; genome sequencing; in utero; mutant; obligate intracellular parasite; pathogen; release of sequestered calcium ion into cytoplasm; research study; response; success

DESCRIPTION (provided by applicant): Toxoplasma gondii is an obligate intracellular parasite known to chronically infect a third of the world's human population. T. gondii infections can lead to severe disease and death in immuno-compromised individuals and significant birth defects when acquired in utero. T. gondii depends on its ability to move into and out of cells to survive and propagate within an infected host. Events required for T. gondii invasion, egress and motility are regulated by calcium signaling processes that include proteins and factors significantly divergent from those of the human host. Thus, inhibition of the calcium signaling pathways that control essential events like invasion and egress constitutes a realistic method to combat this pathogen. Such a strategy requires a better understanding of T. gondii's calcium signaling and accordingly we have taken a forward genetic approach to the identification of calcium signaling proteins. In the last decade we have isolated various mutants with defective responses to artificially induced calcium fluxes. Phenotypic analysis of these mutants has allowed us to learn about various steps leading to the initiation of motility in response to calciu fluxes. Nonetheless, our understanding of these strains and calcium signaling is incomplete, as we have, until recently, been unable to identify the affected genes. The fast pace of development of sequencing techniques has made whole genome sequencing accessible to individual laboratories and thus we have begun revisiting these orphan T. gondii mutants. We have shown the strength of this approach, as well as our ability to harness it, in identifying a mutation in a calcium dependent protein kinase (TgCDPK3) as the cause of the phenotypes in one of these mutants. Currently, two mutants with distinct calcium signaling related phenotypes remain molecularly uncharacterized. Building on our recent success using whole genome sequencing, we propose to sequence the remaining mutants as to identify and begin functional characterization of proteins involved in calcium signaling in T. gondii. Since this study is exploratory and focuses on further analysis of an existing data set, it is ideal for the RO3 fundin mechanism. PUBLIC HEALTH RELEVANCE: One of the most widespread protozoan parasites, Toxoplasma gondii, can lead to severe disease and even death on immunocompromised individuals as well as in the developing fetus. It is the focus of our work to identify the protein involved in signaling the initiation of movement in this parasite. Since inhibiting motility would control this opportunistic infection, our work will lead to the discovery of new drug targets and therapies to combat toxoplasmosis.

描述（由申请人提供）：刚地弓形虫是一种专性细胞内寄生虫，已知慢性感染世界上三分之一的人口。弓形虫感染可导致免疫功能受损的个体严重疾病和死亡，并在子宫内获得严重的出生缺陷。弓形虫依靠其进出细胞的能力在被感染的宿主内存活和繁殖。弓形虫的入侵、出口和运动所需的事件是由钙信号过程调节的，其中包括与人类宿主明显不同的蛋白质和因子。因此，抑制钙信号通路控制基本事件，如入侵和出口，构成了对抗这种病原体的现实方法。这样的策略需要更好地了解弓形虫的钙信号，因此我们已经采取了一种前向遗传方法来鉴定钙信号蛋白。在过去的十年中，我们已经分离出各种对人工诱导的钙通量有缺陷反应的突变体。对这些突变体的表型分析使我们能够了解到在响应钙通量时导致运动性启动的各种步骤。尽管如此，我们对这些菌株和钙信号的理解是不完整的，因为直到最近，我们还无法识别受影响的基因。测序技术的快速发展使得单个实验室可以进行全基因组测序，因此我们开始重新审视这些孤体弓形虫突变体。我们已经证明了这种方法的强度，以及我们利用它的能力，在确定钙依赖性蛋白激酶（TgCDPK3）的突变是这些突变之一的表型的原因。目前，两种具有不同钙信号相关表型的突变体在分子上尚未表征。基于我们最近使用全基因组测序的成功，我们建议对剩余的突变体进行测序，以鉴定并开始对弓形虫钙信号传导相关蛋白进行功能表征。由于本研究是探索性的，并且侧重于对现有数据集的进一步分析，因此它是RO3基金机制的理想选择。