Symbiotic Rickettsia Species as a Model System for the Study of Folate Biosynthes

共生立克次体作为叶酸生物合成研究的模型系统

• 批准号: 8289045
• 负责人: Jianmin Zhong
• 金额: $ 35.35万
• 依托单位: HUMBOLDT STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8289045
• 关键词: Amino Acids; Anabolism; Arthropods; Bacteria; Bacterial Genes; Bioinformatics; Biological Assay; Biological Models; Black-legged Tick; Blood; Bovine Anaplasmosis; Breeding; Cells; Cloning; Complement; Computational algorithm; Computer software; DNA Sequence; Data; Development; Diet; Eukaryotic Cell; Folate; Gene Expression Profile; Genes; Genome; Genomics; Goals; In Vitro; Ixodes; Life; Lyme Disease; Metabolic; Metabolism; Molecular; Nutrient; Open Reading Frames; Pathway interactions; Production; Proteins; Quantitative Reverse Transcriptase PCR; Relative (related person); Research; Rickettsia; Source; Staging; Tick-Borne Diseases; Ticks; United States; Vertebrates; Vitamin B Complex; Vitamins; Work; Writing; base; cofactor; disorder prevention; feeding; fitness; folic acid metabolism; insight; interest; reconstruction

DESCRIPTION (provided by applicant): Nonpathogenic nutrient-providing bacterial endosymbionts have been shown to contribute to their hosts' fitness by supplying them with essential vitamins and amino acids. The metabolic basis for the symbiotic relationship of endosymbionts in ticks has been unknown. Recent metabolic reconstruction, PCR, and DNA sequencing results in our lab have shown that a Rickettsia species in Ixodes pacificus carries genes for the anabolism of cofactors and vitamins. The biosynthetic pathway of folate (vitamin B9) is of particular interest because previous bioinformatic metabolic reconstruction in our lab indicates that the pathway exists in its entirety in the genome of the Rickettsia species. The overall objective of the project is to study bacterial vitamin biosynthetic genes that are essentia for the symbiotic relationship with the tick host. We hypothesize that the endosymbiotic Rickettsia species synthesizes folate, and that the vitamin is utilized by the tick host. The proposed work has three major aspects: 1) Characterization of genes in the folate pathway of the Rickettsia species in Ixodes pacificus using PCR, cloning, and sequencing. 2) Characterization of gene expression patterns of genes in the folate pathway of the Rickettsia species in Ixodes pacificus using quantitative reverse transcriptase PCR (qRT-PCR). 3) Characterization of function of gene products in the folate biosynthetic pathway of the Rickettsia species in Ixodes pacificus by complementation and in vitro enzymatic assay. The research will increase our understanding of the metabolism of folate in ticks, as well as the metabolic basis of symbiotic interactions between the symbiotic Rickettsia species and Ixodes pacificus ticks. Describing the functions of the biosynthetic genes and defining the capacity of the vitamins contributed by the Rickettsia species in Ixodes pacificus, will provide important insights into molecular mechanisms of the endosymbiotic relationship in Ixodes pacificus. PUBLIC HEALTH RELEVANCE: Lyme borreliosis and anaplasmosis are the most common tick-borne diseases in the United States, and prevention of these diseases has become the focus of current tick research. Bacteria that live within eukaryotic cells have allowed their hosts to acquire metabolic capabilities otherwise unavailable. These bacterial partners contribute to their hosts' fitness by providing essential nutrients such as vitamins and amino acids. Arthropods that feed primarily or entirely on blood are virtually a breeding ground for these types of partnerships because vertebrate blood rarely contains sufficient quantities of essential B-vitamins and amino acids. Therefore, the diet of blood feeding arthropods is complemented with bacterially synthesized vitamins (2, 5, 76). The sole diet of most ticks is blood from vertebrates, but the sources of B-vitamins for Ixodes pacificus ticks are unknown. The recent sequencing of the Ixodes scapularis genome revealed genomic sequences of the tick's bacterial partner. Using freely available computer software, as well as computer algorithms written in our lab, we have discovered all six genes involved in the vitamin B9 biosynthetic pathway. The pathway is from a Rickettsia species in Ixodes pacificus, the West Coast relative of Ixodes scapularis. In this proposal, we hypothesize that the Rickettsia species, living within Ixodes pacificus cells, produces essential B9 vitamin that is not synthesized by Ixodes pacificus. We will determine if all genes in the B9 vitamin pathway of the Rickettsia species are actively transcribed in Ixodes pacificus ticks, and if these bacterial genes encode proteins that synthesize vitamin B9. The proposed studies will offer an ideal model system to study the impact of bacterial endosymbionts on the metabolism of ticks and more specifically, the effect of the Rickettsia endosymbiont on Ixodes pacificus fitness.

描述(申请人提供)：非致病的营养提供细菌内共生体已被证明有助于其宿主的健康，通过提供必要的维生素和氨基酸。硬蜱体内共生菌共生关系的代谢基础尚不清楚。我们实验室最近的代谢重建、聚合酶链式反应和DNA测序结果表明，太平洋硬蜱中的一种立克次体携带辅因子和维生素合成代谢的基因。叶酸(维生素B9)的生物合成途径特别令人感兴趣，因为我们实验室先前的生物信息代谢重构表明，该途径完整地存在于立克次体的基因组中。该项目的总体目标是研究细菌维生素生物合成基因，这些基因是与扁虱宿主共生关系的必需品。我们假设内共生立克次体合成叶酸，维生素被壁虱宿主利用。这项工作主要包括三个方面：1)利用聚合酶链式反应、克隆和测序技术对太平洋硬蜱中立克次体的叶酸途径基因进行了鉴定。2)应用定量逆转录聚合酶链式反应(qRT-PCR)分析太平洋硬蜱叶酸途径相关基因的表达模式。3)通过互补法和体外酶法鉴定基因产物在太平洋硬蜱叶酸生物合成途径中的功能。这项研究将加深我们对硬蜱体内叶酸代谢的了解，以及共生立克次体与太平洋硬蜱共生相互作用的代谢基础。描述生物合成基因的功能，确定太平洋硬蜱中立克次体所提供的维生素的能力，将为揭示和平硬蜱内生共生关系的分子机制提供重要的信息。 公共卫生相关性：莱姆疏螺旋体病和无浆体病是美国最常见的硬蜱传播疾病，预防这些疾病已成为当前硬蜱研究的重点。生活在真核细胞内的细菌允许它们的宿主 获得原本无法获得的新陈代谢能力。这些细菌伙伴通过提供维生素和氨基酸等基本营养物质来促进宿主的健康。主要或完全以血液为食的节肢动物实际上是这些类型的繁殖地。 这是因为脊椎动物的血液中很少含有足够数量的必需B族维生素和氨基酸。因此，食血节肢动物的饮食中补充了细菌合成的维生素(2，5，76)。大多数扁虱的唯一食物是脊椎动物的血液， 但太平洋硬蜱的B族维生素来源尚不清楚。最近对肩部硬蜱基因组的测序揭示了扁虱的细菌伙伴的基因组序列。使用免费提供的计算机软件，以及我们实验室编写的计算机算法，我们已经发现了参与维生素B9生物合成途径的所有六个基因。这条途径来自太平洋硬蜱中的一种立克次体，它是肩部硬蜱的西海岸近亲。在这项提议中，我们假设生活在太平洋硬蜱细胞内的立克次体物种产生了必要的B9维生素，而这种维生素不是由和平硬蜱合成的。我们将确定是否Rickettsia物种的B9维生素途径中的所有基因都在太平洋硬蜱中活跃地转录，以及这些细菌基因是否编码合成维生素B9的蛋白质。这些研究将为研究细菌内共生菌对硬蜱新陈代谢的影响，更具体地说，研究内共生立克次体对太平洋硬蜱适合性的影响提供一个理想的模型系统。

项目成果

The Rickettsia Endosymbiont of Ixodes pacificus Contains All the Genes of De Novo Folate Biosynthesis.

• DOI: 10.1371/journal.pone.0144552
• 发表时间: 2015
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Hunter DJ;Torkelson JL;Bodnar J;Mortazavi B;Laurent T;Deason J;Thephavongsa K;Zhong J
• 通讯作者: Zhong J