Oxidative Stress Enhances AP-1 and NF-κB-Mediated Regulation of β2-Glycoprotein I Gene Expression in Hepatoma Cells

beta(2)-Glycoprotein I (beta(2)-GPI), also known as apolipoprotein H, is a plasma glycoprotein with poorly defined gene regulation. The aim of this study was to clarify the role of oxidative stress in beta(2)-GPI gene regulation and determine the essential transcription element regulating beta(2)-GPI expression. We demonstrate that expression of beta(2)-GPI at the protein and mRNA levels was significantly elevated in Huh7 and HepG2 cells treated with 100 mu M hydrogen peroxide (H2O2). To address the transcriptional mechanism of H2O2-mediated beta(2)-GPI gene regulation, several promoter constructs were cloned and characterized by deletion assays. A region spanning from -2141 to -1419 (relative to the transcription start site), which contains two activator protein-1 (AP-1) sites (AP 1-2 and AP1-3) and one nuclear factor-kappaB (NF-kappa B) site was found to be the main target site for up-regulation of beta(2)-GP1 promoter activity by oxidative stress. In addition, we found that H2O2 stimulation enhanced the nuclear translocation of AP-1 and NF-kappa B subunits. Using an electrophoretic mobility shift assay, it was confirmed that nuclear protein binding to the API-2, AP1-3, and NF-kappa B sites was increased in Huh7 cells treated with H2O2. Knockdown of the c-Jun, c-Fos, p6.5, and p50 genes using small interfering RNAs (siRNAs) further confirmed that AP-1 and NF-kappa B play an essential role in the H2O2-induced beta(2)-GPI expression. Overall, these findings provide new insight suggesting that multiple cis-elements in the beta(2)-GPI promoter work cooperatively to regulate beta(2)-GPI expression in cells under oxidative stress. J. Cell. Biochem. 111: 988-998, 2010. zolo Wiley-Liss, Inc.