Regulation of airway epithelial cell NF-κB-dependent gene expression by protein kinase Cδ¹

Airway epithelial cells synthesize proinflammatory molecules such as IL-8, GM-CSF, RANTES, and ICAM-1, the expression of which is increased in the airways of patients with asthma. We investigated the regulation of these NF-κB-dependent genes by the novel protein kinase C (PKC) isoform PKCδ in 16HBE14o-human airway epithelial cells, focusing on IL-8 expression. Transient transfection with the constitutively active catalytic subunit of PKCδ (PKCδ-CAT), and treatment with bryostatin 1, an activator of PKCδ, each increased transcription from the IL-8 promoter, whereas overexpression of PKCε had minor effects. Expression of a dominant negative PKCδ mutant (PKCδ-KR) or pretreatment of cells with rottlerin, a chemical PKCδ inhibitor, attenuated TNF-α- and phorbol ester-induced transcription from the IL-8 promoter. Bryostatin 1 treatment increased IL-8 protein abundance in primary airway epithelial cells. Selective activation of PKCδ by bryostatin also activated NF-κB, as evidenced by p65 RelA and p50 NF-κB1 binding to DNA, NF-κB trans-activation, and IκB degradation. The sufficiency of PKCδ to induce NF-κB nuclear translocation and binding to DNA was confirmed in a 16HBE14o-cell line inducibly expressing PKCδ-CAT under the tet-off system. Deletion of the NF-κB response element severely attenuated PKCδ-induced IL-8 promoter activity. Finally, PKCδCAT induced transcription from the GM-CSF, RANTES, and ICAM-1 promoters. Together these data suggest that PKCδ plays a key role in the regulation of airway epithelial cell NF-κB-dependent gene expression.