Kidney-specific expression of human organic cation transporter 2 (OCT2/SLC22A2) is regulated by DNA methylation
Kidney-specific expression of human organic cation transporter 2 (OCT2/SLC22A2) is regulated by DNA methylation
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DOI：
10.1152/ajprenal.90257.2008
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发表时间：
2008-07-01
影响因子：
4.2
通讯作者：
Inui, Ken-ichi
中科院分区：
文献类型：
--
作者：
Aoki, Masayo;Terada, Tomohiro;Inui, Ken-ichi
人体有机阳离子转运蛋白2 （OCT2/SLC22A2）在肾脏中特异性表达，在肾脏中阳离子化合物的分泌中起关键作用。OCT2的组织表达和膜定位与其底物药物的组织分布、药理作用和/或不良反应密切相关。然而，OCT2肾脏特异性表达的分子机制尚未阐明。因此，在本研究中，我们使用人体组织样本检测了OCT2基因启动子区域DNA甲基化对其组织特异性表达的贡献。利用从肾脏和肝脏中提取的人类基因组DNA，通过亚硫酸盐测序研究了OCT2近端启动子区和OCT1（肝脏特异性有机阳离子转运体）的体内甲基化状态。OCT2近端启动子中的所有CpG位点在肝脏中都是高甲基化的，而在肾脏中是低甲基化的。另一方面，OCT1的启动子区域在肾脏和肝脏中都出现了高甲基化。在基础转录因子上游刺激因子（USF） 1的结合位点E-box的CpG位点，OCT2启动子的甲基化水平尤其低。在体外，OCT2近端启动子的甲基化显著降低了转录活性，电泳迁移率转移实验表明，E-box的甲基化抑制了USF1的结合。这些结果表明，人类OCT2的肾特异性表达受近端启动子区甲基化调控，干扰USF1的反激活。
Human organic cation transporter 2 (OCT2/SLC22A2), which is specifically expressed in the kidney, plays critical roles in the renal secretion of cationic compounds. Tissue expression and membrane localization of OCT2 are closely related to the tissue distribution, pharmacological effects, and/or adverse effects of its substrate drugs. However, the molecular mechanisms underlying the kidney-specific expression of OCT2 have not been elucidated. In the present study, therefore, we examined the contribution of DNA methylation of the promoter region for the OCT2 gene to its tissue-specific expression using human tissue samples. In vivo methylation status of the proximal promoter region of OCT2 and that of OCT1, a liver-specific organic cation transporter, were investigated by bisulfite sequencing using human genomic DNA extracted from the kidney and liver. All CpG sites in the OCT2 proximal promoter were hypermethylated in the liver, while hypomethylated in the kidney. On the other hand, the promoter region of OCT1 was hypermethylated in both the kidney and liver. The level of methylation of the OCT2 promoter was especially low at the CpG site in the E-box, the binding site of the basal transcription factor upstream stimulating factor (USF) 1. In vitro methylation of the OCT2 proximal promoter dramatically reduced the transcriptional activity, and an electrophoretic mobility shift assay showed that methylation at the E-box inhibited the binding of USF1. These results indicate that kidney-specific expression of human OCT2 is regulated by methylation of the proximal promoter region, interfering with the transactivation by USF1.