Characterization of activating signal cointegrator-2 as a novel transcriptional coactivator of the xenobiotic nuclear receptor constitutive androstane receptor
Characterization of activating signal cointegrator-2 as a novel transcriptional coactivator of the xenobiotic nuclear receptor constitutive androstane receptor
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DOI：
10.1210/me.2005-0066
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发表时间：
2005-07-01
影响因子：
--
通讯作者：
Kim, SW
中科院分区：
文献类型：
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作者：
Choi, E;Lee, S;Kim, SW
激活信号协整子-2 （activated signal cointegrator-2, ASC-2）是最近分离到的一种转录辅助激活蛋白，用于多种不同的转录因子，包括核受体超家族的许多成员。在本报告中，我们证明ASC-2也可以作为外生核受体组成型雄甾烷受体（CAR）的辅激活剂。首先，在CV-1和HeLa细胞中，共转染ASC-2可增强CAR的转录激活。相反，在稳定表达针对ASC-2的特异性小干扰RNA的HepG2细胞中，CAR转激活显著受损。与这些结果一致，染色质免疫沉淀实验显示ASC-2以配体依赖的方式被募集到已知的CAR靶基因上。其次，CAR特异性地与ASC-2的第一个LXXLL基序相互作用，这些相互作用被CAR激动剂1,4-二苯[2（3,5-二氯吡啶氧基）]苯刺激，并被CAR逆激动剂雄甾醇抑制，表明该基序可能介导ASC-2和CAR在体内的相互作用。为了支持这一观点，包含第一个LXXLL基序的ASC-2片段DN1抑制CAR转激活，并共表达ASC-2，但其他LXXLL型共激活因子（如甲状腺激素受体相关蛋白220）不能逆转这种抑制。最后，最近发现CAR在影响对乙酰氨基酚引起的严重肝损伤中起关键作用。有趣的是，表达DN1的转基因小鼠对对乙酰氨基酚诱导的肝毒性具有抗性，并且在这些转基因小鼠中，一系列已知CAR靶基因的表达被特异性抑制。综上所述，这些结果强烈表明ASC-2是一个真正的外源核受体CAR的共激活因子，并介导CAR在体内的特异性外源反应。
Activating signal cointegrator-2 (ASC-2) is a recently isolated transcriptional coactivator protein for a variety of different transcription factors, including many members of the nuclear receptor superfamily. In this report, we demonstrate that ASC-2 also serves as a coactivator of the xenobiotic nuclear receptor constitutive androstane receptor ( CAR). First, transcriptional activation by CAR was enhanced by cotransfected ASC-2 in CV-1 and HeLa cells. In contrast, CAR transactivation was significantly impaired in HepG2 cells stably expressing specific small interfering RNA directed against ASC-2. Consistent with these results, chromatin immunoprecipitation experiments revealed that ASC-2 is recruited to the known CAR target genes in a ligand-dependent manner. Secondly, CAR specifically interacted with the first LXXLL motif of ASC-2, and these interactions were stimulated by CAR agonist 1,4-bis[ 2( 3,5-dichloropyridyloxy)] benzene and repressed by CAR inverse agonist androstanol, suggesting that this motif may mediate the interactions of ASC-2 and CAR in vivo. In support of this idea, DN1, a fragment of ASC-2 encompassing the first LXXLL motif, suppressed CAR transactivation, and coexpressed ASC-2 but not other LXXLL-type coactivators such as thyroid hormone receptor-associated protein 220 reversed this repression. Finally, CAR was recently found to play a pivotal role in effecting the severe acetaminophen-induced liver damage. Interestingly, transgenic mice expressing DN1 were resistant to the acetaminophen-induced hepatotoxicity and expression of a series of the known CAR target genes was specifically repressed in these transgenic mice. Taken together, these results strongly suggest that ASC-2 is a bona fide coactivator of the xenobiotic nuclear receptor CAR and mediate the specific xenobiotic response by CAR in vivo.