Role of the carboxy-terminal transactivation domain and active transcription in the ligand-induced and ligand-independent degradation of the mouse Ahb-1 receptor.
Role of the carboxy-terminal transactivation domain and active transcription in the ligand-induced and ligand-independent degradation of the mouse Ahb-1 receptor.
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羧基末端反式激活结构域和活性转录在配体诱导和配体独立的小鼠 Ahb-1 受体降解中的作用。
DOI：
10.1016/j.bcp.2005.09.006
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发表时间：
2005
影响因子：
5.8
通讯作者：
Dougherty,EdwardJ
中科院分区：
文献类型：
--
作者：
Pollenz,RichardS;Popat,Jesal;Dougherty,EdwardJ
为探讨转录激活结构域（transactivation domain，ERK）、DNA结合和转录对AH受体（AH receptor，AHR）降解的重要性，用放线菌素D（actinomycin D，AD）或放线菌酮（cycloheximide，CHX）预处理Hepa-1细胞，并暴露于2，3，7，8-四氯二苯并-对-二恶英（tetrachlorodibenzo-p-dioxin，TCDD）。AD或CHX不影响AHR的核定位或DNA结合，但抑制配体诱导的降解。与此相反，AD或CHX不能抑制格尔德霉素（GA）诱导的AHR降解。为了评估COOH末端的密码子在AHR降解中的作用，将终止密码子置于Ahb−1AHR编码区的核苷酸1501和1921处，以产生AHR 500和AHR 640。产生稳定的细胞系并暴露于TCDD。表达AHR 500的细胞不诱导CYP 1A 1蛋白，但表现出显著的AHR 500降解。表达AHR 640的细胞诱导的CYP 1A 1蛋白水平为表达野生型AHR的细胞的50%，并表现出显著的AHR 640降解。重要的是，AD和CHX不抑制TCDD诱导的AHR 500和AHR 640的降解，并且与表达野生型AHR的细胞相比，这些受体显示出更快的配体诱导降解谱。TCDD暴露于芳烃受体核转位蛋白（ARNT）减少的Hepa-1细胞，显示配体诱导的AHR降解，而AD未阻断。然而，AD抑制TCDD诱导的降解时，ARNT表达恢复。这些结果表明，存在多种机制的配体和GA诱导的降解的AHR，并表明，配体诱导的降解可以在两种机制之间切换，这取决于一个功能性的酶的存在和DNA的结合。
To assess the importance of transactivation domains (TAD), DNA binding and transcription on the degradation of the AH receptor (AHR), Hepa-1 cells were pre-treated with actinomycin D (AD) or cycloheximide (CHX) and exposed to 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). AD or CHX did not affect nuclear localization or DNA binding of the AHR but inhibited ligand-induced degradation. In contrast, AD or CHX did not inhibit geldanamycin (GA) induced degradation of the AHR. To assess the role of the COOH-terminal TAD in AHR degradation, stop codons were placed at nucleotide 1501 and 1921 of the Ahb−1AHR coding region to generate AHR500and AHR640. Stable cell lines were generated and exposed to TCDD. Cells expressing AHR500did not induce CYP1A1 protein, but exhibited significant degradation of AHR500. Cells expressing AHR640induced CYP1A1 protein to 50% of the level of cells expressing wild type AHR and exhibited significant degradation of AHR640. Importantly, AD and CHX did not inhibit the TCDD-induced degradation of either AHR500and AHR640and these receptors showed a more rapid profile of ligand-induced degradation compared to cells expressing wild type AHR. TCDD exposure to Hepa-1 cells with reduced aryl hydrocarbon receptor nuclear translocator (ARNT), showed ligand-induced degradation of the AHR that was not blocked by AD. However, AD inhibited TCDD-induced degradation when ARNT expression was restored. These results show that multiple mechanisms exist for the ligand and GA-induced degradation of the AHR and suggest that ligand-induced degradation can switch between two mechanisms depending on the presence of a functional TAD and the binding to DNA.