請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/49548
標題: | let-7微小核醣核酸調控核苷酸二磷酸激酶NME4基因表現 Expression of nucleoside diphosphate kinase NME4 is regulated by the let-7 microRNA |
作者: | Chun-Ming Hsu 徐君明 |
指導教授: | 詹世鵬 |
關鍵字: | 微小核醣核酸,核?酸二磷酸激?,粒線體, let-7,NME4,mitochondria, |
出版年 : | 2016 |
學位: | 碩士 |
摘要: | 微小核醣核酸(miRNA)是一種新穎的微小非編碼核醣核酸,它在後轉錄的層次上藉由和目標基因的三端不轉譯區以鹼基配對的方式來調控各種下游基因的表現。某些微小核醣核酸異常的表現會扮演著腫瘤抑制因子的角色,例如let-7,若不正常表現將會導致人類發育異常及腫瘤生成。在我們的報導中NME4是一個高可性度的let-7的目標基因,在人類胚胎腎細胞、肝癌細胞中加入過量的let-7模擬物會使NME4蛋白質有減少的趨勢,相反地,在用let-7的反義股抑制住let-7後使NME4蛋白質有增加的趨勢。接著我們將NME4的三端不轉譯區選殖到帶有冷光酵素基因的載體中,並且測量在細胞中加入了let-7模擬物及抑制物後冷光活性的變化,我們發現了let-7模擬物會降低帶有NME4三端不轉譯區質體的螢光表現而用let-7反義股將let-7抑制住則會提高螢光表現,另外,在TargetScan軟體預測NME4三端不轉譯區上的let-7結合位進行突變卻能使此現象消失,因此綜合以上我們認為NME4會受到let-7的調控。 MicroRNAs (miRNAs) are an emerging class of small non-coding RNAs that regulate expression of various downstream genes at the post-transcriptional level, achieved through partial base-pairing between miRNAs and the target sequences at the 3’UTRs of mRNAs. Aberrant expression of certain miRNAs serving as tumor suppressors, such as let-7, leads to abnormal development and conduces to progression of human cancers. Here we report that the non-metastatic cells 4 (NME4) is a “high-confidence” let-7 target. The protein levels of NME4, examined by western blots, were down-regulated when excess amount of let-7 mimics were introduced into HEK293, Huh7 or HepG2 cells. By contrast, inhibition of let-7 by introducing antisense inhibitors increased the NME4 protein levels. We cloned the NME4 3’UTR into a luciferase vector and measured the luciferase activity of lysates prepared from the cells transfected with let-7 mimics or inhibitors. We found that let-7 mimics reduced the luciferase activity and let-7 inhibitors showed an opposite effect. Mutations of the putative let-7 target site at the NME4 3’UTR, predicted by the TargetScan miRNA target prediction software, eliminated the effects on the luciferase activity. Thus, we propose that NME4 is regulated by the let-7 miRNA. The biological implication of the let-7-NME4 regulation awaits further investigation. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/49548 |
DOI: | 10.6342/NTU201602961 |
全文授權: | 有償授權 |
顯示於系所單位: | 微生物學科所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-105-1.pdf 目前未授權公開取用 | 3.21 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。