p11 expression in human bronchial epithelial cells is increased by nitric oxide in a cGMP-dependent pathway involving protein kinase G activation.

The effect of nitric oxide on p11 expression was studied in an immortalized human bronchial epithelial cell line (BEAS-2B cells). Three nitric oxide donors were used: spermine NONOate (SP), (+/-)-S-nitroso-N-acetylpenicillamine (SNAP), and S-nitrosoglutathione (SNOG). All three nitric oxide donors had similar effects resulting in dose-dependent and time-dependent accumulation of p11 protein and an increase of steady-state p11 mRNA. Studies using a reporter gene containing the region from -1499 to +89 of the p11 promoter demonstrated an increase in transcriptional activity after stimulation with NO donors for 4 h. These effects were abolished at the promoter and protein level using protein kinase G inhibitors (KT5823 and R(p)-8-pCPT-cGMPS). Incubation of transfected cells with a cell permeable cGMP analogue (8-Br-cGMP) resulted in a dose-related increase of promoter activity. An electrophoretic mobility shift assay of nuclear proteins extracted from BEAS-2B cells identified an AP-1 site located at -82 to -77 of the p11 promoter region as an NO- and cGMP- dependent response element. These data were confirmed using a c-jun dominant negative mutant vector and a c-jun expression plasmid. Therefore, we conclude that nitric oxide-induced p11 expression in human bronchial epithelial cells is mediated at least in part through increased binding of activator protein one to the p11 promoter.