Cytochromes P450 (CYPs) catalyze a great number of metabolic reactions that have profound effects on the biological activities of xenobiotics and endobiotics. In this study, we aimed to characterize rhythmic expressions of drug-metabolizing CYPs using synchronized hepatoma cells, and to investigate the potential roles of cis-elements of circadian clock system (E-box, D-box and RevRE or RORE) in generating the rhythms.

Serum was used to synchronize circadian cycles and to induce circadian gene expression in cultured hepatoma cells (HepRG and HepG2 cells). Regulation of CYP genes by circadian clock components was investigated by performing luciferase reporter, overexpression and knockdown experiments. mRNA and protein expression were determined by qPCR and Western blotting assays, respectively.

Of ten major drug-metabolizing CYP genes, six are rhythmically expressed (CYP1A2, 2B6, 2C8, 2D6, 2E1 and 3A4), whereas other four are non-rhythmic (CYP1B1, 2A6, 2C9 and 2C19).

The E-box binding protein BMAL1 directly controls the rhythmic expression of CYP1A2. Rhythmic expressions of CYP2E1 and CYP3A4 are generated via both E-box and D-box elements. The RevRE binding protein REV-ERBα contributes to rhythmic oscillations in CYP2B6 and CYP2C8.

In conclusion, rhythmic expressions of five human CYPs (CYP1A2, 2B6, 2C8, 2E1 and 3A4) are generated and regulated by E-box-, D-box-, and/or RevRE-acting clock components. Our findings may have implications for understanding chronopharmacokinetic events in humans.

细胞色素P450（CYP）催化大量的代谢反应，这些反应对异种生物和内生生物的生物活性产生深远的影响。在这项研究中，我们旨在表征使用同步化肝癌细胞的药物代谢性CYP的节律性表达，并研究昼夜节律时钟系统（E-box，D-box和RevRE或RORE）的顺式元素在体内产生的潜在作用。节奏。

血清用于同步昼夜节律周期并诱导培养的肝癌细胞（HepRG和HepG2细胞）中的昼夜节律基因表达。通过进行荧光素酶报道基因，过表达和敲低实验研究了昼夜节律对CYP基因的调控。mRNA和蛋白质表达分别通过qPCR和Western印迹测定来确定。

在十种主要的药物代谢CYP基因中，有节奏地表达了六个（CYP1A2、2B6、2C8、2D6、2E1和3A4），而其他四个则是非有节奏的（CYP1B1、2A6、2C9和2C19）。

E-box结合蛋白BMAL1直接控制CYP1A2的节律表达。CYP2E1和CYP3A4的节奏表达是通过E-box和D-box元素生成的。RevRE结合蛋白REV-ERBα有助于CYP2B6和CYP2C8的节律性振荡。

总之，五种人CYP（CYP1A2、2B6、2C8、2E1和3A4）的节律性表达是由E-box-，D-box-和/或RevRE作用时钟组件产生和调节的。我们的发现可能对理解人类体内的药代动力学事件有影响。