NF-κB mediates the basal expression of ATX. A. Alignment of the ATX promoter region 2000 nt upstream of the transcription start site from 9 eutherian mammals. Two highly conservative consensus sequences for NF-κB binding sites are identified. B. Starved Hep3B cells were treated with parthenolide (0, 2.5 or 5 μM) for 16 hours. C. ATX expression in stable cell line Hep3B-IκBαSR which NF-κB activity was blocked by mutant IκBα (IκBαSR). Vector infected cell line Hep3B-pQCXIN served as control; IκBαSR indicates Hep3B-IκBαSR cells; pQCXIN indicates Hep3B-pQCXIN cells. ATX mRNA expression was determined by qRT-PCR (top panel in B, C), Values are the mean ± SD of three experiments. ** P < 0.01, * P < 0.05. ATX protein expression was determined by immunoblot (IB) (Bottom panel in B, C). β-actin expression was used as a loading control.