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B-Dependent Gene Expression by Protein Kinase C
1
* Department of Pediatrics, and
Ben May Institute for Cancer Research, University of Chicago, Chicago, IL 60637;
Herbert Irving Comprehensive Cancer Center, College of Physicians and Surgeons, Columbia University, New York, NY 10032; and the
Department of Medicine, University of Texas Medical Branch, Galveston, TX 77555
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-
B-dependent genes by the novel protein kinase C (PKC) isoform PKC
in 16HBE14o- human airway epithelial cells, focusing on IL-8 expression. Transient transfection with the constitutively active catalytic subunit of PKC (PKC-CAT), and treatment with bryostatin 1, an activator of PKC, each increased transcription from the IL-8 promoter, whereas overexpression of PKC
had minor effects. Expression of a dominant negative PKC mutant (PKC-KR) or pretreatment of cells with rottlerin, a chemical PKC inhibitor, attenuated TNF-
- 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 PKC by bryostatin also activated NF-B, as evidenced by p65 RelA and p50 NF-B1 binding to DNA, NF-B trans-activation, and IB degradation. The sufficiency of PKC to induce NF-B nuclear translocation and binding to DNA was confirmed in a 16HBE14o- cell line inducibly expressing PKC-CAT under the tet-off system. Deletion of the NF-B response element severely attenuated PKC-induced IL-8 promoter activity. Finally, PKC-CAT induced transcription from the GM-CSF, RANTES, and ICAM-1 promoters. Together these data suggest that PKC plays a key role in the regulation of airway epithelial cell NF-B-dependent gene expression.
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