請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/68529
標題: | 探討第八型岩藻醣轉移酶藉活化表皮生長因子受體而調控角質細胞增生 Fucosyltransferase 8 (FUT8) Mediates Epidermal Growth Factor Receptor (EGFR) Activation to Regulate Keratinocyte Proliferation |
作者: | Yu-Tien Huang 黃于恬 |
指導教授: | 李永凌 |
關鍵字: | 乾蘚,第八型岩藻醣轉移?,醣基化,表皮細胞生長因子受體,細胞增生,角質細胞, Psoriasis,Fucosyltransferase 8,Glycosylation,Epidermis growth factor receptor,Proliferation,Keratinocyte, |
出版年 : | 2017 |
學位: | 碩士 |
摘要: | 研究目的及方法:
乾蘚是一個常見的慢性遺傳性皮膚疾病,其主要特徵為表皮增生分化出現不正常。表皮細胞生長因子受體為表皮細胞膜上與細胞增生高度相關的蛋白,此受體及其主要配體已經被發現在乾蘚病灶處大量表達。在哺乳動物體內,第八型岩藻醣轉移酶是唯一可以對蛋白質進行核心岩藻醣基化的酵素。在過去的實驗當中,我們證實在人類角質細胞中的第八型岩藻醣轉移酶可以促進細胞的增生。表皮細胞生長因子受體為一醣基化蛋白,並已證實在多種癌症中受到第八型岩藻醣轉移酶調控。因此本研究旨在討論人類角質細胞中經由第八型岩藻醣轉移酶調控的表皮細胞生長因子受體活化反應。 首先經由臨床資料與檢體分析乾蘚病灶與非病灶中第八型岩藻醣轉移酶的表現量,並計算其與臨床嚴重程度的相關性。其次經由醣蛋白體學技術找出第八型岩藻醣轉移酶在其變異細胞株中的關鍵醣蛋白。並經由免疫染色等方式呈現第八型岩藻醣轉移酶變異細胞株中與正常角質細胞相異的表皮細胞生長因子受體其本身與下游磷酸化訊息傳遞鍊,二聚體構成,配體結合力以及表面受體內部化等等活化反應。 結果與結論: 免疫染色結果顯示乾蘚病灶相較於非病灶中有較高的第八型岩藻醣轉移酶表現量,且其表現量比值與臨床嚴重程度成正向關係。由醣蛋白體學技術找出第八型岩藻醣轉移酶變異細胞株中,表皮細胞生長因子受體的核心岩藻醣受到調控。在生長因子配體刺激下,表皮細胞生長因子受體其本身與下游磷酸化訊息傳遞鍊,二聚體構成,配體結合力以及表面受體內部化等等活化反應在第八型岩藻醣轉移酶弱化的細胞株中都呈現較正常細胞株低下的反應,而這些影響則間接造成角質細胞增生異常。 Background: Psoriasis is a common, heritable, and chronic inflammatory skin disease. Epidermal hyperplasia is directly caused by keratinocyte hyperproliferation. Epidermis growth factor receptor (EGFR), a transmembrane receptor highly related with cell proliferation, and its ligands were reported to be up-regulated in psoriasis lesion skin. Fucosyltransferase 8 (FUT8) is the only enzyme to catalyze 1,6-fucosylation in mammal and has been observed to regulated cell proliferation. Since EGFR is a core fucosylated protein, we aim to investigate the regulatory role of FUT8 on EGFR activation. Method: FUT8 expression in psoriasis was examined with immunofluorescence. In vitro studies were conducted under lentiviral transduction of FUT8 overexpress and knockdown on normal human keratinocyte cell line (HaCaT). Core fucosylated glycoproteins were isolated by LC MS/MS with SILAC labeling. Immunoprecipitation and immunoblotting were used to investigate EGFR core fucosylation, phosphorylation and downstream signaling. BS3 cross linker was used to observe EGFR dimerization. Alkaline phosphatase report system was used to investigate EGF binding affinity. Alexa-488 conjugated EGF was used to observe EGFR internalization. Results: FUT8 was up-regulated in psoriasis lesion skin. Knockdown of FUT8 suppresses EGFR core fucosylation, EGFR phosphorylation, MAPK signaling, EGFR dimerization, EGF binding affinity and EGFR internalization in human keratinocyte cell line (HaCaT). |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/68529 |
DOI: | 10.6342/NTU201704080 |
全文授權: | 有償授權 |
顯示於系所單位: | 流行病學與預防醫學研究所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-106-1.pdf 目前未授權公開取用 | 1.8 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。