Mycobacterium bovis bacillus Calmette-Guerin (BCG)-induced CXCL8 production is mediated through PKCα-dependent activation of the IKKαβ signaling pathway in epithelial cells

Upon contact with airway epithelial cells, mycobacteria activate several signal transduction events that are required for induction of NF-kappa B-dependent chemokine gene expression. However, downstream signaling pathways, especially that of Ca2+-dependent protein kinase C alpha (PKC alpha), and in particular, the identity of the IKK alpha beta signal pathway for CXCL8 secretion in Mycobacterium bovis BCG-induced epithelial cells are still unknown. In this study, we demonstrated that the phosphoinositide-phospholipase C (PI-PLC) downstream signaling pathway is involved in M. bovis BCG-induced CXCL8 release, since A549 cells pretreated with U73122, a PI-PLC inhibitor, inhibited CXCL8 release, whereas U73343 the inactive analog had no effect. In addition, our results demonstrated that M. bovis BCG-induced CXCL8 production by A549 cells was significantly blocked by using neomycin (another well-described inhibitor of PI-PLC with a different mechanism of action), Ro-32-0432 and Ro-31-8220 (two PKC alpha inhibitors), PP1 and PP2 (two potent and selective inhibitors of the Src-family tyrosine kinases), and Bay 11-7082 (an I kappa B phosphorylation inhibitor). We also demonstrated that M bovis BCG can rapidly induce translocation of PKC alpha from the cytosol to the membrane, and that treatment of cells with M. bovis BCG caused time-dependent increases in phosphorylation of c-Src at tyrosine 416. Finally, our studies revealed that M. bovis BCG induced the association of c-Src and IKK alpha beta during the interaction of PKC alpha and IKK alpha beta. Altogether, these results represent the first evidence to date suggesting that M. bovis BCG activates the PI-PLC/PKC alpha/c-Src/IKK alpha beta signaling pathway to induce CXCL8 release in human epithelial cells. (c) 2007 Elsevier Inc. All rights reserved.