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  • 學位論文

陰道鞭毛蟲滋養體及阿米巴體比較蛋白質體學與轉錄體學之研究

Comparative proteomics and transcriptomics of Trichomonas vaginalis trophozoite and amoeboid stages.

指導教授 : 梁有志

摘要


陰道鞭毛蟲症是目前世界上最普遍經由性行為感染的疾病。陰道鞭毛蟲基因體全長約170 MB,其含有六萬個基因,是目前已知原蟲當中最多的。由鞭毛體型態轉變成吸附的阿米巴形態對陰道的感染是很重要的步驟。在這個研究中我們利用了比較轉錄體學與蛋白質體學的系統分析方法針對陰道鞭毛蟲滋養體與阿米巴體的分子調控做探討。我們首先建立了陰道鞭毛蟲滋養體的標準二維電泳蛋白圖譜,再利用比較二維電泳結合質譜的方法找出了49個表現量有差異的蛋白。其中在阿米巴體中具有抗氧化壓力活性的蛋白表現量是增加的,而醣類代謝及細胞骨架蛋白表現量則是下降的。轉錄體學則利用大規模表現序列標籤(ESTs)的定序來探討基因表現。由滋養體與阿米巴體的cDNA libraries 分析超過兩萬個ESTs,並依據它們的功能做分類。我們也選取了在阿米巴體時期基因表現量大於5倍(116個基因)及10倍(32個基因)的基因做分析。有趣的是,與壓力有關的基因在阿米巴體時期被大量的表現,表示阿米巴體時期是在壓力的狀態之下。這也暗示這些抗氧化壓力的蛋白在阿米巴體時期扮演重要的角色,且阿米巴體時期對陰道鞭毛蟲來說並不是最佳的生理狀態。這些由轉錄體學及蛋白質體學分析所得到的發現希望可以提供對陰道鞭毛蟲症提供新的觀點並且對此寄生蟲與宿主之間的作用有更基礎的了解。

並列摘要


Trichomoniasis caused by Trichomonas vaginalis is the most common sexual transmitted infection in the world. The 170-MB genome of this protozoan contains 60,000 genes, the largest number of genes ever identified in protozoan. The morphological transformation from a flagellated form to an adherent amoeboid form is crucial to the establishment of infection in vagina. In this study, we established the reference two-dimensional gel electrophoresis (2-DGE) map of the trophozoite stage. The molecular response in the amoeboid and trophozoite stages of T .vaginalis was elucidated by using a systemic comparative transcriptomics and proteomics approach. We identified 49 differentially expressed proteins by 2-DGE and MALDI-TOF MS. Among them, stress proteins with antioxidant properties were up-regulated whereas carbohydrate metabolism and cytoskeletal proteins were down-regulated in the amoeboid form. Transcriptomics analysis at gene expression level was performed by using large scale expressed sequence tags (ESTs) sequencing. More than 20,000 ESTs from trophozoite and amoeboid cDNA libraries were clustered and classified according to their biological functions. We also summarized the genes which expressed 5-fold (116 genes) and 10-fold (32 genes) higher in the amoeboid stage. Interestingly, most of these significantly expressed genes in the amoeboid form are stress-related genes, indicating that the amoeboid stage is under stress. This implied that these antioxidant proteins have important roles in the amoeboid stage and the pathogenic amoeboid stage is not the optimum physiological condition for T. vaginalis. These findings derived from proteome and transcriptome analysis are expected to provide new insights into the pathogenesis of trichomoniasis and will facilitate a more fundamental understanding of the host-parasite interplay.

參考文獻


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