The role of polyphosphate in Toxoplasma gondii

聚磷酸盐在弓形虫中的作用

• 批准号: 10681078
• 负责人: Silvia N Moreno
• 金额: $ 21.28万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-13 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10681078
• 关键词: AIDS-Related Opportunistic Infections; Address; Alkalinization; Bacteria; Binding; Bioinformatics; Calcium; Catalogs; Catalytic Domain; Cells; Cellular Metabolic Process; Cellular Stress; Client; Complex; Cryptosporidiosis; Cryptosporidium; Cyst; Cytolysis; Cytosol; Data; Diatoms; Disease; Electrostatics; Energy-Generating Resources; Enzymes; Eukaryota; Experimental Designs; Future; Gene Expression; Genome; Heat Stress Disorders; Homologous Gene; Human; Immune response; Immunocompromised Host; Infection; Ions; Knowledge; Link; Liquid substance; Lytic Phase; Malaria; Mammalian Cell; Modification; Molecular Chaperones; Morbidity - disease rate; Names; Orthologous Gene; Parasites; Pathogenesis; Pathogenicity; Pathway interactions; Persons; Phase; Plasmodium; Play; Polymers; Polyphosphates; Polyps; Population; Post-Translational Protein Processing; Process; Proteins; Proteome; Regulation; Role; Rupture; Saccharomyces cerevisiae; Signal Transduction; System; Testing; Tissues; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Vacuole; Vertebrate Biology; Vertebrates; Virulence; Virulence Factors; Yeasts; acute infection; biological adaptation to stress; chronic infection; cohort; forgetting; fungus; human disease; inorganic phosphate; knock-down; member; metal chelator; mortality; mutant; novel; response; scaffold; validation studies

PROJECT SUMMARY/ABSTRACT Apicomplexan parasites cause persistent mortality and morbidity worldwide through diseases like malaria, toxoplasmosis, and cryptosporidiosis. The phylum member Toxoplasma gondii alone infects approximately one third of the world population. The pathogenesis of T. gondii is reliant on its capacity to replicate within host cells and spread to other host cells. The acute infection is ultimately controlled by the host immune response and the parasite establishes a chronic infection characterized by the presence of tissue cysts containing slow replicating bradyzoites. Tissue cyst rupture releases bradyzoites that convert into tachyzoites that replicate fast and if the host is immunocompromised, could cause severe and even fatal tissue damage. We propose to study how polyphosphate (polyP), a “forgotten molecule”, exerts its role in pathogenicity and virulence of T. gondii. PolyP is a ubiquitous polymer of three to hundreds of phosphate residues linked by high-energy phosphoanhydride bonds and can reach millimolar levels in protozoan parasites, while the concentration in host cells is at the micromolar level. T. gondii stores polyP at molar levels in acidocalcisomes which are hydrolyzed upon alkalinization followed by release of Ca2+ into the cytosol. It is puzzling how little is known about the function of polyP in the T. gondii infection cycle and/or virulence. However, characterizing how polyP functions will impact our knowledge of the mechanism by which T. gondii causes disease. Both in bacteria and eukaryotes polyP is involved in a large number of diverse and apparently unrelated functions. However, these apparently unrelated functions may be the result of polyP’s ability to interact with proteins either electrostatically or covalently and we propose to test this for T. gondii. Our hypothesis is that polyP exerts its essential function in the lytic cycle of T. gondii through its interaction with proteins and/or their covalent modification by polyphosphorylation. We propose two strategies to determine how polyP exerts its role in virulence. To define candidate proteins that interact with polyP we will use Thermal Proteome Profiling and expose intact parasites and/or lysates to heat stress to underscore the role of polyP as a protein-like chaperone. We will also use a bioinformatics approach to discover potential proteins that are covalently modified by polyP. We propose to use polyP mutants (that do not make polyP) already available for the characterization of the role of polyP interaction with target proteins. The synthesis of polyP represents a novel target as it is essential, and the enzymes are absent in the mammalian host. The proteins involved in polyP synthesis are conserved in other apicomplexan parasites. The polyP target proteins will be specific for T. gondii virulence pathways and could have orthologs in other apicomplexans. Additionally, findings from this project could have wider impact for vertebrate biology since the mammalian polyP synthesis machinery is unknown and our findings may uncover novel functions and proteins.

项目总结/摘要 顶复门寄生虫通过疟疾等疾病在世界范围内引起持续的死亡率和发病率， 弓形体病和隐孢子虫病。门成员弓形虫单独感染大约 世界人口的三分之一。对T.弓形虫依赖于它在体内复制的能力， 宿主细胞并扩散到其他宿主细胞。急性感染最终由宿主免疫控制 反应和寄生虫建立一个慢性感染的特点是存在组织囊肿 含有缓慢复制的缓殖子组织囊肿破裂释放缓殖子转化为速殖子 如果宿主免疫力低下，这种病毒会快速复制，并可能导致严重甚至致命的组织损伤。 我们建议研究多磷酸盐（polyP），一种“被遗忘的分子”，如何发挥其致病作用 和毒力。刚地。PolyP是一种普遍存在的聚合物，由三到数百个磷酸残基连接 通过高能磷酸酐键，在原生动物寄生虫中可以达到毫摩尔水平，而 在宿主细胞中的浓度为微摩尔水平。T.弓形虫在酸钙体中以摩尔水平储存polyP 其在碱化后水解，随后将Ca 2+释放到胞质溶胶中。令人费解的是 关于T中polyP的功能是已知的。弓形虫感染周期和/或毒力。然而，表征 polyP如何发挥作用将影响我们对T.弓形虫引起疾病。 在细菌和真核生物中，polyP都参与了大量不同且显然无关的蛋白质合成。 功能协调发展的然而，这些明显不相关的功能可能是polyP与细胞相互作用的能力的结果。 蛋白质静电或共价，我们建议测试T。刚地。我们的假设是 polyP在T细胞裂解周期中发挥重要作用。通过与蛋白质的相互作用 和/或它们通过多磷酸化的共价修饰。 我们提出了两种策略，以确定聚P如何发挥其作用的毒力。为了确定候选蛋白质 我们将使用热蛋白质组分析并将完整的寄生虫和/或裂解物暴露于 热应激以强调聚P作为蛋白质样伴侣的作用。我们还将使用生物信息学 方法来发现潜在的蛋白质，被聚P共价修饰。我们建议使用polyP 突变体（不产生polyP）已经可用于表征polyP与 靶蛋白。聚P的合成代表了一个新的靶点，因为它是必需的，而酶却不存在 在哺乳动物宿主中。参与polyP合成的蛋白质在其他顶复门中是保守的 寄生虫聚P靶蛋白将对T.弓形虫毒力途径，可能有直系同源物 在其他apicomplexans。此外，该项目的发现可能对脊椎动物生物学产生更广泛的影响 由于哺乳动物polyP的合成机制尚不清楚，我们的发现可能会揭示新的功能， 和蛋白质。