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標題: | 探討熱帶念珠菌中六個轉錄因子對於菌絲生成及生物膜發育的調控功能 Insight into the function of six transcription factors in regulation of filamentation and biofilm development in Candida tropicalis |
作者: | Chien-Jui Hou 侯健睿 |
指導教授: | 林晉玄 |
關鍵字: | 熱帶念珠菌,轉錄因子,調控網路,生物膜發育,菌絲生成, Candida tropicalis,transcription factors,regulatory network,biofilm development,filamentation, |
出版年 : | 2017 |
學位: | 碩士 |
摘要: | 熱帶念珠菌 (Candida tropicalis) 是一種伺機性致病真菌,能造成表面黏膜感染以及可致命的菌血症 (candidemia)。為適應環境,熱帶念珠菌能在不同的型態間轉換,如酵母菌 (yeast)、真菌絲 (hyphae)、偽菌絲 (pseudohyphae) 等。菌絲生成能幫助C. tropicalis侵入宿主表皮黏膜組織,同時也對生物膜發育相當重要。生物膜不但加強菌株抗藥性、增加治療難度,也是病人癒後復發的重要因素。過去研究發現,白色念珠菌 (C. albicans) 中由六個轉錄因子Bcr1、Brg1、Efg1、Ndt80、Rob1、Tec1組成的調控網路能調節生物膜發育。本研究欲探討六個C. tropicalis中六個轉錄因子對於生物膜發育及菌絲生成的影響。實驗發現牛血清刺激下野生株MYA 3404能生成菌絲 (> 20%),而野生株ATCC 34139無法生成 (< 1%)。因此,本研究利用MYA 3404作為標準菌株進行突變株構築。研究顯示,突變株的生物膜生成均有缺失,與野生株比較後生物膜重量顯著下降 (WT: 5.16 mg; bcr1∆: 0.50 mg; brg1∆: 2.58 mg; efg1∆: 0.02 mg; ndt80∆: 0.42 mg; rob1∆: 3.45 mg; tec1∆: 2.51 mg)。以牛血清進行菌絲生成測試,結果顯示突變株菌絲比例與野生株亦有顯著差異 (WT: 39.17%; bcr1∆: 23.54%; brg1∆: 18.49%; efg1∆: 0.53%; ndt80∆: 60.53%; rob1∆: 28.59%; tec1∆: 11.06%) 。有趣的是, ndt80∆ 菌絲生成比例大幅度上升。進一步以Spider-Glu 不同培養基觀察菌絲生成,除了rob1∆菌絲比例與野生株沒有差異外,其餘突變株結果與上述相似 (WT: 4.23%; bcr1∆: 0.86%; brg1∆: 0.88%; efg1∆: 1.89%; ndt80∆: 24.41%; rob1∆: 7.29%; tec1∆: 1.41%)。結果顯示熱帶念珠菌Bcr1、Brg1、Efg1、Ndt80、Rob1、Tec1對生物膜生成的功能具保守性且為正向調控。然而,與白色念珠菌不同的是,Ndt80負向調控熱帶念珠菌的菌絲生成。 Candida tropicalis is an opportunistic human fungal pathogen that can cause mucosal infections and life-threatening candidemia. To adapt to different environmental niches, C. tropicalis could switch between yeast, true hyphae and pseudohyphae. Hyphae not only help the invasion to host cells, but also play critical role in biofilm development, thereby hampering antifungal drug treatment and recurrence. Previous report showed that six transcription factors, including Bcr1, Brg1, Efg1, Ndt80, Rob1, and Tec1, form a core transcriptional circuit to control biofilms. Thus, the aim of this study is to investigate the six C. tropicalis homologous transcriptional genes in regulation of hyphae formation and biofilm development. Under serum induction, the wildtype stran MYA 3404 (> 20%) forms filaments while wildtype strain ATCC 34139 (< 1%) do not. Thus, the following experiments and mutant constuctions will use the reference strain MYA 3404. Comparison with the wildtype strain, mutant strains of each gene caused a significant reduction in biofilms (WT: 5.16 mg; bcr1∆: 0.50 mg; brg1∆: 2.58 mg; efg1∆: 0.02 mg; ndt80∆: 0.42 mg; rob1∆: 3.45 mg; tec1∆: 2.51 mg). During the response to bovine serum, hyphal formation between each mutant and the wildtype strain also resulted in significant difference (YL477: 39.17%; bcr1∆: 23.54%; brg1∆: 18.49%; efg1∆: 0.53%; ndt80∆: 60.53%; rob1∆: 28.59%; tec1∆: 11.06%). Interestingly, hyphae in the C. tropicalis ndt80∆ highly increased. Under Spider-Glu culture condition, similar results were observed, except rob1∆ in which it showed comparable filamentation percentages to those of the wildtype strain (YL477: 4.23%; bcr1∆: 0.86%; brg1∆: 0.88%; efg1∆: 1.89%; ndt80∆: 24.41%; rob1∆: 7.29%; tec1∆: 1.41%). These data suggest that Bcr1, Brg1, Efg1, Ndt80, Rob1 and Tec1 in C. tropicalis play conserved roles in biofilm formation. However, different to the C. albicans Ndt80, Ndt80 in C. tropicalis negatively regulates hyphal formation. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/59939 |
DOI: | 10.6342/NTU201700204 |
全文授權: | 有償授權 |
顯示於系所單位: | 生化科技學系 |
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