PPM1A Phosphatase is Involved in Regulating Pregnane Xenobiotic Receptor Mediated Cytochrome P450 3A4 Gene Expression

Abstract

The liver is the most important organ of drug metabolism and plays a major role in the detoxification of both endobiotics and xenobiotics. The process of drug metabolism is undertaken in three distinct phases. Cytochromes p450s (CYPs), enzymes belonging to the first phase of drug metabolism, are vital to drug metabolism. In particular, cytochrome p450 3A4 (CYP3A4), which metabolizes 60% of FDA approved drugs, plays a crucial role in drug metabolism. The human pregnane xenobiotic receptor (hPXR), a ligand-dependent orphan nuclear receptor, is a major transcription factor that regulates the expression of key drug-metabolizing enzymes, including CYP3A4. Variations in the expression of hPXR-mediated CYP3A4 in liver can alter therapeutic response to a variety of drugs and may lead to potential adverse drug interactions. However, molecular mechanisms of hPXR-mediated CYP3A4 expression are not fully understood. We sought to determine whether Mg2+/Mn2+-dependent phosphatase 1A (PPM1A) regulates hPXR-mediated CYP3A4 expression in liver hepatocytes. PPM1A was found to be coimmunoprecipitated with hPXR. Genetic or pharmacologic activation of PPM1A led to a significant increase in hPXR transactivation of CYP3A4 promoter activity. In contrast, knockdown of endogenous PPM1A not only attenuated hPXR transactivation, but also increased proliferation of HepG2 cells. This suggests that PPM1A expression levels regulate hPXR, and that PPM1A expression and hPXR activity are regulated in a proliferation-dependent manner. Indeed, PPM1A expression and hPXR transactivation were significantly reduced in sub-confluent HepG2 cells compared to confluent HepG2 cells, suggesting that both PPM1A expression and hPXR-mediated CYP3A4 expression may be downregulated in proliferating livers. Elevated PPM1A levels led to attenuation of hPXR inhibition by tumor necrosis factor-alpha and cyclin-dependent kinase 2 which are known to be upregulated and essential during liver regeneration. In mice regenerating livers, similar to sub-confluent HepG2 cells, expression of both PPM1A and the mouse PXR target gene cyp3a11 (human ortholog of CYP3A4) was found to be downregulated. Together, our results show that PPM1A is essential for PXR function and can positively regulate PXR activity by counteracting PXR inhibitory signaling pathways that play a major role in liver regeneration. These results implicate a novel role for PPM1A in regulating hPXR-mediated CYP3A4 expression in liver hepatocytes, and may explain a mechanism for CYP3A repression in regenerating livers.