Activated Sterol Regulatory Element-Binding Protein-2 Suppresses Hepatocyte Nuclear Factor-4-Mediated Cyp3a11 Expression in Mouse Liver
Activated Sterol Regulatory Element-Binding Protein-2 Suppresses Hepatocyte Nuclear Factor-4-Mediated Cyp3a11 Expression in Mouse Liver
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DOI：
10.1124/mol.110.068577
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发表时间：
2011-01-01
影响因子：
3.6
通讯作者：
Yamazoe, Yasushi
中科院分区：
文献类型：
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作者：
Inoue, Shin-ichi;Yoshinari, Kouichi;Yamazoe, Yasushi
甾醇调节元件结合蛋白-2 （SREBP-2）是胆固醇稳态的关键转录因子。最近的研究表明，主要的药物代谢酶CYP3A酶与胆固醇代谢有关。在本研究中，我们研究了SREBP-2参与肝脏Cyp3a11表达的可能性。给小鼠喂食低胆固醇饮食（LCD）激活了肝脏SREBP-2，同时减弱了肝脏Cyp3a11的表达。在LCD中添加胆固醇可以逆转这些现象。在报告基因实验中，组成型活性SREBP-2的过表达降低了Cyp3a11报告基因在- 1581到- 1570区域的活性。该区域含有一个假定的肝细胞核因子-4 α (HNF-4 α)结合基序，在体外结合实验中，HNF-4 α与基序结合，而不是SREBP-2。随着该基序的突变或缺失，在报告基因分析中，依赖srebp -2的Cyp3a11表达抑制消失。在拉下实验和共免疫沉淀实验中，SREBP-2通过其反式激活结构域与过氧化物酶体增殖体激活受体γ共激活因子-1 α (PGC-1 α)结合，抑制了HNF-4 α和PGC-1 α之间的相互作用。缺乏反活化结构域的突变体SREBP-2始终未能降低Cyp3a11报告基因的活性。此外，PGC-1 α过表达减轻了srebp -2介导的Cyp3a11报告基因活性的降低。最后，染色质免疫沉淀实验表明，PGC-1 α与Cyp3a11启动子的结合程度在小鼠肝脏中通过lcd喂养而降低。综上所述，在胆固醇摄入量降低的小鼠肝脏中，活化的SREBP-2与PGC-1 α相互作用。这导致PGC-1 α在Cyp3a11启动子上募集到HNF-4 α减少，随后Cyp3a11表达下调。
Sterol regulatory element-binding protein-2 (SREBP-2) is a key transcription factor for the cholesterol homeostasis. Recent studies have suggested the association of CYP3A enzymes, major drug-metabolizing enzymes, with cholesterol metabolism. In the present study, we have investigated a possible involvement of SREBP-2 in hepatic Cyp3a11 expression. Feeding a low-cholesterol diet (LCD) to mice activated hepatic SREBP-2 whereas it attenuated hepatic Cyp3a11 expression. These phenomena were reversed by cholesterol supplementation to LCD. In reporter assays, the overexpression of constitutively active SREBP-2 reduced Cyp3a11 reporter activity through the region from - 1581 to - 1570 of Cyp3a11. This region contained a putative hepatocyte nuclear factor-4 alpha (HNF-4 alpha) binding motif, and HNF-4 alpha, but not SREBP-2, bound to the motif in in vitro binding assays. With the mutation or deletion of this motif, the SREBP-2-dependent suppression of Cyp3a11 expression disappeared in reporter assays. In pull-down assays and coimmunoprecipitation assays, SREBP-2 bound to peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1 alpha), a major coactivator for HNF-4 alpha, via its transactivation domain and inhibited the interaction between HNF-4 alpha and PGC-1 alpha in vitro. A mutant SREBP-2 lacking the transactivation domain consistently failed to reduce Cyp3a11 reporter activity. Furthermore, PGC-1 alpha overexpression relieved the SREBP-2-mediated reduction of Cyp3a11 reporter activity. Finally, chromatin immunoprecipitation assays demonstrated that the extent of PGC-1 alpha binding to the Cyp3a11 promoter was reduced by LCD-feeding in mouse livers. In conclusion, activated SREBP-2 interacts with PGC-1 alpha in mouse livers at reduced cholesterol intake. This results in the reduced PGC-1 alpha recruitment to HNF-4 alpha on the Cyp3a11 promoter and the subsequent down-regulation of Cyp3a11 expression.