Protein Phosphatase 2CβI Regulates Human Pregnane X Receptor-Mediated CYP3A4 Gene Expression in HepG2 Liver Carcinoma Cells

The human pregnane X receptor ( hPXR) regulates the expression of CYP3A4, which plays a vital role in hepatic drug metabolism and has considerably reduced expression levels in proliferating hepatocytes. We have recently shown that cyclin-dependent kinase 2 (CDK2) negatively regulates hPXR-mediated CYP3A4 gene expression. CDK2 can be dephosphorylated and inactivated by protein phosphatase type 2C beta isoform long (PP2C beta I), a unique phosphatase that was originally cloned from human liver. In this study, we sought to determine whether PP2C beta I is involved in regulating hPXR's transactivation activity and whether PP2C beta I affects CDK2 regulation of this activity in HepG2 liver carcinoma cells. In transactivation assays, transiently coexpressed PP2C beta I significantly enhanced the hPXR-mediated CYP3A4 promoter activity and decreased the inhibitory effect of CDK2 on hPXR transactivation activity. In addition, shRNA-mediated down-regulation of endogenous PP2C beta I promoted cell proliferation, inhibited the interaction of hPXR with steroid receptor coactivator-1, and attenuated the hPXR transcriptional activity. The levels of PP2C beta I did not affect hPXR expression. Our results show for the first time that PP2C beta I is essential for hPXR activity and can positively regulate this activity by counteracting the inhibitory effect of CDK2. Our results implicate a novel and important role for PP2C beta I in regulating hPXR activity and CYP3A4 expression by inhibiting or desensitizing signaling pathways that negatively regulate the function of pregnane X receptor in liver cells and are consistent with the notion that both the activity of hPXR and the expression of CYP3A4 are regulated in a cell cycle-dependent and cell proliferation-dependent manner.