Nuclear receptor coactivator 6 mediates the synergistic activation of human cytochrome P-4502C9 by the constitutive androstane receptor and hepatic nuclear factor-4α
Nuclear receptor coactivator 6 mediates the synergistic activation of human cytochrome P-4502C9 by the constitutive androstane receptor and hepatic nuclear factor-4α
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DOI：
10.1124/mol.108.048983
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发表时间：
2008-09-01
影响因子：
3.6
通讯作者：
Goldstein, Joyce A.
中科院分区：
文献类型：
--
作者：
Surapureddi, Sailesh;Rana, Ritu;Goldstein, Joyce A.
核受体辅激活因子 6 (NCOA6) 也称为 PRIP/RAP250/ASC-2，锚定辅因子的稳态复合物，并充当某些核受体的转录辅激活因子。这是第一项将 NCOA6 鉴定为肝核因子 4 α (HNF4 α) 相互作用蛋白的研究。 CYP2C9 是一种重要的酶，可代谢常用的治疗药物和重要的内源性化合物。我们之前已经证明，组成型雄甾烷受体（CAR）（一种异生素感应受体）通过与远端位点结合上调 CYP2C9 启动子，而 HNF4 α 通过近端位点转录上调 CYP2C9。我们证明了 CAR 和 HNF4 α 之间配体增强的协同串扰。我们认为 NCOA6 对于这种串扰的潜在机制至关重要。在本研究中，使用酵母双杂交筛选和 GST Pull-down 测定，NCOA6 被鉴定为 HNF4 α 相互作用蛋白。此外，我们还发现 NCOA6、CAR 和其他辅激活因子是 HepG2 细胞中与 HNF4 α 相关的大型辅因子复合物的一部分。尽管 NCOA6 与 CAR 的相互作用具体是通过 NCOA6 的第一个 LXXLL 基序，但这两个 LXXLL 基序都参与其与 HNF4 α 的相互作用。 NCOA6 的沉默消除了 CYP2C9 启动子的协同激活以及 CAR-HNF4 α 对 CYP2C9 基因的协同诱导。染色质免疫沉淀分析显示，NCOA6 可以拉低 CYP2C9 启动子的近端 HNF4 α 和远端 CAR 结合位点，并为其他辅因子的招募提供基础。我们得出结论，共激活剂 NCOA6 介导 CAR 和 HNF4 α 协同激活 CYP2C9 基因的机制。
Nuclear receptor coactivator 6 (NCOA6) also known as PRIP/RAP250/ASC-2 anchors a steady-state complex of cofactors and function as a transcriptional coactivator for certain nuclear receptors. This is the first study to identify NCOA6 as a hepatic nuclear factor 4 alpha (HNF4 alpha)-interacting protein. CYP2C9 is an important enzyme that metabolizes both commonly used therapeutic drugs and important endogenous compounds. We have shown previously that constitutive androstane receptor (CAR) (a xenobiotic-sensing receptor) up-regulates the CYP2C9 promoter through binding to a distal site, whereas HNF4 alpha transcriptionally up-regulates CYP2C9 via proximal sites. We demonstrate ligand-enhanced synergistic cross-talk between CAR and HNF4 alpha. We identify NCOA6 as crucial to the underlying mechanism of this cross-talk. NCOA6 was identified as an HNF4 alpha-interacting protein in this study using a yeast two-hybrid screen and GST pull-down assays. Furthermore, we identified NCOA6, CAR, and other coactivators as part of a mega complex of cofactors associated with HNF4 alpha in HepG2 cells. Although the interaction of NCOA6 with CAR is specifically through the first LXXLL motif of NCOA6, both LXXLL motifs are involved in its interaction with HNF4 alpha. Silencing of NCOA6 abrogated the synergistic activation of the CYP2C9 promoter and the synergistic induction of the CYP2C9 gene by CAR-HNF4 alpha. Chromatin immunoprecipitation analysis revealed that NCOA6 can pull down both the proximal HNF4 alpha and distal CAR binding sites of the CYP2C9 promoter and provides the basis for the recruitment of other cofactors. We conclude that the coactivator NCOA6 mediates the mechanism of the synergistic activation of the CYP2C9 gene by CAR and HNF4 alpha.