https://scholars.lib.ntu.edu.tw/handle/123456789/163429
標題: | 組蛋白去乙醯基轉移酵素阻礙劑抑制腎上腺皮質細胞內類固醇轉錄因子的表現 Inhibitors of Histone Deacetylase Down-regulate the Expression of Steroidogenic Factor 1 in Adrenocorticol Cells |
作者: | 賴政分 lai, Cheng-Fen |
關鍵字: | 類;固醇轉錄;因子;孤核受體;腎上腺皮質腫瘤細胞;組蛋白去乙醯基轉移酵素抑制劑;乙醯化;膽固醇側鏈分切酵素;上游刺激因子一及二;histone acetylatransferase;P450 side chain cleavage enzyme;the cholesterol side-chain cleavage enzyme;steroidogenic factor 1;trichostatin A | 公開日期: | 2005 | 摘要: | 類固醇轉錄因子(Steroidogenic Factor1, SF-1)是孤核受體(orphan nuclear receptor)的一種,能調控許多類固醇生成酵素基因的表現,對腦下垂體、下視丘、腎上腺及性線的發育及分化非常重要。在小鼠與人的腎上腺皮質腫瘤細胞內(Y1 and H295 cells),使用組蛋白去乙醯基轉移酵素抑制劑 ,如Trichostatin A和Sodium butyrate,能增加Histone 4的乙醯化 (Acetylation),卻減少SF-1與其下游膽固醇側鏈分切酵素(P450scc)的表現,這個抑制現象與劑量和作用時間成正比。報導基因表現分析的實驗中,TSA抑制了SF-1啟動子的活性;TSA能增加外源性SF-1蛋白質的表現,但減少內源性SF-1蛋白質的表現量。表示HDAC 抑制劑是透過SF-1啟動子來控制基因轉錄的表現。許多文獻指出,SF-1啟動子上面,具有一個重要的調控序列,E-box,與其結合的兩個主要蛋白質為上游刺激因子一及二 (Upstream Stimulatory Factor 1 and USF2)。膠體電泳位移分析 (EMSA) 結果表示,HDAC 抑制劑同時減少SF-1啟動子上調控單位(E box and CCAAT box)與其轉錄因子間的結合,也減少了USF1和USF2蛋白質的表現量。TSA抑制了P450scc及USF2的mRNA,但卻增加了USF1的mRNA;此結果與減少的USF1蛋白質相矛盾,故使用HDAC 抑制劑可能會影響特定蛋白質的穩定性並加速其降解。 The orphan nuclear receptor steroidogenic factor 1 (SF-1, A4BP, or NR5A1) is a key transcription factor that regulates the expression of many steroidogenic enzymes. It has also been demonstrated to play an important role in the development and differentiation of hypothalamus, pituitary, adrenal glands, and gonads. Being such an important factor, the regulation of SF-1 expression is highly investigated. We used inhibitors of histone deacetylases which generally activate gene expression and used as anticancer drugs such as trichostatin A and sodium butyrate. Histone deacetylase inhibitors although increased the acetylation of Histone 4, reduced the protein content of SF1 and cytochrome P450 side-chain cleavage enzyme, a SF-1 controlled gene, in a dose and time-dependent manner in adrenocorticol cells, Y1 and H295 cells. In a reporter assay, TSA reduced the SF-1 promoter activity. In Y1 stable clone, which expresses SF-1-HA driven by CMV promoter, TSA increased exogenous SF-1-HA protein, however, reduced endogenous SF-1 protein. Both results indicate that SF-1 is transcriptionally down-regulated by these inhibitors. TSA reduced binding of E-box and CCAAT box binding factor to SF-1 promoter in Electorphoretic Mobility Shift Assay. The E-box binding proteins, USF1 (upstream stimulatory factor 1) and USF2 were down-regulated by TSA as well as SF-1. These results suggest that TSA-inhibited expression of SF-1 may provide insight to uncover the regulatory mechanism of SF-1 gene. Interesting, we found that TSA treatment increases mRNA of SCC and USF2 but reduces USF1 mRNA in Quantitative-Real-Time PCR. Increased USF1 mRNA and reduced USF1 protein content after TSA treatment provides a new association of protein stability and acetylation. |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/59975 | 其他識別: | en-US |
顯示於: | 獸醫學系 |
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ntu-94-R92629015-1.pdf | 23.31 kB | Adobe PDF | 檢視/開啟 |
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