2017-08-012024-05-14https://scholars.lib.ntu.edu.tw/handle/123456789/659304摘要：陰道滴蟲感染引起的陰道滴蟲症是最常見非病毒源引起之性傳染疾病，更可能增加人類免疫不全病毒的傳播，並發現可能與子宮頸癌與前列腺癌病程發展相關。早先實驗全細胞蛋白質體分析以及膠體內蛋白質體分析法已發現鐵離子可能活化陰道滴蟲去乙醯酶(TvHDAC1)的Ser391 位點磷酸化，並改變細胞核中組織蛋白乙醯化程度。進一步利用改良細胞核蛋白質體學分析法發現多個陰道滴蟲的組織蛋白去乙醯酶(TvHDACs)以及組織蛋白乙醯轉化酶(TvHATs)被磷酸化，顯示鐵離子刺激可能誘發陰道滴蟲大規模表徵遺傳調控。本計畫將針對此些新發現的去乙醯酶與乙醯轉化酶近行生化特性分析，並將其野生型與突變蛋白過量表現於細胞中以進行功能性分析。到目前為止，我們已發現TvHDAC1 的組織蛋白受質，在本研究計畫將以膠體內蛋白質體學分析法確認新發現的陰道滴蟲TvHDACs 與TvHATs 其受質為何，並進一步利用西方轉漬與免疫螢光染色偵測確認。此外，我們將利用成熟的高通量定序法分析染色質體免疫沉澱產物，以解析細胞受鐵刺激後TvHDACs 與TvHATs 或各式乙醯化組織蛋白在陰道滴蟲基因體上的DNA 結合模式，以分析結合區域之基因分佈。此外，我們也發現高吸附力的陰道滴蟲細胞株，組織蛋白H3K9 乙醯化程度遠高於低吸附力的細胞株，此現在顯示表徵遺傳機制亦可能參與調控陰道滴蟲致病力表現。本計畫所提出全新的概念，為陰道滴蟲鐵誘發表徵遺傳調控研究提供重要訊息，並有利於原蟲領域之細胞學發展。<br> Abstract: Trichomonas vaginalis causes trichomoniasis, is the most common non-viral sexually transmitteddisease, and infection increases the risk of HIV transmission, cervical cancer and prostate cancerdevelopment. In our previous whole-cell phosphoproteomic and gel-based proteomic analysis, iron wasfound to induce TvHDAC1 Ser391 phosphorylation and change histone acetylation status. By using improvednuclear protein phosphoproteomic approach, phosphorylation sites specific to multiple TvHDACs andTvHATs were detected to be activated in T. vaginalis upon sudden iron overloading, strongly suggesting thatiron may activate tremendous epigenetic regulation to control histone acetylation status in this parasite. Inthis proposal, we will characterize biochemical nature of those newly-identified TvHDACs and TvHATs, andoverexpression of wild type or dominant negative mutants will be introduced for further functional dissection.So far, the substrates of TvHDAC1 have been found, we proposed to identify the histone substrates of eachTvHDAC and TvHAT by a gel-based proteomic approach followed by validation with immunofluorescenceassay or Western Blotting detection. Moreover, we will map those gene targets of TvHADCs and TvHATs orhistone acetylation in T. vaginalis genome, by a well-established chromatin immunoprecipitation coupledwith next generation sequencing to explore those genes in response to iron overloading. Moreover, histoneacetylation status was found to high level in a high-adherent isolate of T. vaginalis, supposing that epigeneticis very likely involved in regulation of T. vaginalis pathogenicity. Our study will provide pioneeringunderstanding in epigenetic mechanism, and beneficial to scientific community of T. vaginalis陰道滴蟲組織蛋白去乙醯酶組織蛋白乙醯轉化酶鐵離子表徵遺傳控基因轉錄訊 息傳導致病力Trichomonas vaginalisHistone deacetylaseHistone acetyltransferaseIronEpigenetic regulationGene transcriptionSignal transductionPathogenicity