Choline phosphotransferase-dependent phospholipid synthesis in Treponema

描述（由申请人提供）：密螺旋体属螺旋体中存在的磷脂酰胆碱（PtdCho）生物合成的独特cdp -胆碱途径可能是这些生物共生/致病生活方式的结果和贡献者，反映了它们与真核宿主的共同进化。我们假设，T. denticola PtdCho合成的最后一个酶步骤（1,2-二酰基甘油胆碱磷酸转移酶[CPT]）是由TDE0021编码的，这是一个明显的真核起源基因。在许多真核生物中，该酶也具有合成磷脂酰乙醇胺（PtdEtn）所需的1,2-二酰基甘油乙醇胺磷酸转移酶（EPT）活性。在T. denticola中，当PtdCho合成的第一步被阻断时，PtdEtn（通过一种尚未确定的途径合成）就会上调。我们假设T. denticola TDE0021编码CPT活性，也可能具有EPT活性，从而使其成为磷脂合成的关键酶，并成为开发针对一系列粘膜和性病密螺旋体病的特异性抗密螺旋体药物的有吸引力的靶点。目的1：表征T. denticola合成PtdCho所需的CPT活性。我们将通过(A)构建和表征一个等基因的TDE0021突变体，以及(B)将一个酿酒酵母CPT/EPT突变体与T. denticola TDE0021 DNA进行互补，来确定T. denticola TDE0021是否编码CPT活性。目的2：研究PtdEtn在牙齿田鼠中的合成和表达。我们将(A)鉴定和表征齿形T. PtdEtn合成途径，以及(B)表征在确定的ptdcho缺陷齿形T.突变体中PtdEtn水平升高的机制。在完成本项目的两个目标后，我们将在了解密螺旋体属（包括粘膜和性病病原体）独特磷脂合成途径的遗传成分和机制方面取得重大进展。所获得的知识将显著增强对螺旋体膜生物学的理解，将为继续研究磷脂环境在微生物-宿主相互作用中的作用提供基础，并可能为开发新的治疗药物提供基础。公共卫生相关性：该项目将描述密螺旋体属中独特磷脂合成途径的遗传成分和机制，其中包括粘膜和性病病原体。这将有助于了解膜磷脂环境在微生物-宿主相互作用中的作用，并可能为开发新的治疗药物提供基础。

DESCRIPTION (provided by applicant): The unique CDP-choline pathway for phosphatidylcholine (PtdCho) biosynthesis present in spirochetes of the genus Treponema is likely both a consequence of and a contributor to the commensal/pathogenic lifestyles of these organisms, reflecting their coevolution with eukaryotic hosts. We hypothesize that the final enzyme step in T. denticola PtdCho synthesis (1,2- diacylglycerol choline phosphotransferase [CPT]) is encoded by TDE0021, a gene of apparent eukaryotic origin. In many eukaryotes, this enzyme also has 1,2-diacylglycerol ethanolamine phosphotransferase (EPT) activity required for phosphatidylethanolamine (PtdEtn) synthesis. In T. denticola, PtdEtn (synthesized by an as-yet unidentified pathway) is upregulated when the first step in PtdCho synthesis is blocked. We hypothesize that T. denticola TDE0021 encodes CPT activity and may also have EPT activity, thus making it the key enzyme in phospholipid synthesis and an attractive target for development of specific anti-Treponema agents against a range of mucosal and venereal treponematoses. This application addresses the following areas: Aim 1: To characterize the CPT activity of T. denticola required for synthesis of PtdCho. We will determine if T. denticola TDE0021 encodes CPT activity by (A) construction and characterization of an isogenic TDE0021 mutant and (B) complementation of a Saccharomyces cerevisiae CPT/EPT mutant with T. denticola TDE0021 DNA. Aim 2: To characterize synthesis and expression of PtdEtn in T. denticola. We will (A) identify and characterize the T. denticola PtdEtn synthesis pathway, and (B) characterize the mechanism(s) responsible for increased levels of PtdEtn in a defined PtdCho-deficient T. denticola mutant. Upon completion of the two Aims of this project, we will have made significant progress toward understanding the genetic components and mechanisms of the unique phospholipid synthesis pathway in the genus Treponema, which includes both mucosal and venereal pathogens. The knowledge gained will significantly enhance understanding of spirochete membrane biology, will provide a basis for continued studies of the role of the phospholipid environment on microbe-host interactions, and may provide the basis for development of novel therapeutic agents. PUBLIC HEALTH RELEVANCE: This project will characterize genetic components and mechanisms comprising the unique phospholipid synthesis pathways in the genus Treponema, which includes both mucosal and venereal pathogens. This will contribute to understanding of the role of the membrane phospholipid environment on microbe-host interactions, and may provide a basis for development of novel therapeutic agents.