PKC zeta participates in activation of inflammatory response induced by enteropathogenic E. coli.
We showed previously that enteropathogenic Escherichia coli (EPEC) infection of intestinal epithelial cells induces inflammation by activating NF-kappa B and upregulating IL-8 expression. We also reported that extracellular signal-regulated kinases (ERKs) participate in EPEC-induced NF-kappa B activation but that other signaling molecules such as PKC zeta may be involved. The aim of this study was to determine whether PKC zeta is activated by EPEC and to investigate whether it also plays a role in EPEC-associated inflammation. EPEC infection induced the translocation of PKC zeta from the cytosol to the membrane and its activation as determined by kinase activity assays. Inhibition of PKC zeta by the pharmacological inhibitor rottlerin, the inhibitory myristoylated PKC zeta pseudosubstrate (MYR-PKC zeta-PS), or transient expression of a nonfunctional PKC zeta significantly suppressed EPEC-induced I kappa B alpha phosphorylation. Although PKC zeta can activate ERK, MYR-PKC zeta-PS had no effect on EPEC-induced stimulation of this pathway, suggesting that they are independent events. PKC zeta can regulate NF-kappa B activation by interacting with and activating I kappa B kinase (IKK). Coimmunoprecipitation studies showed that the association of PKC zeta and IKK increased threefold 60 min after infection. Kinase activity assays using immunoprecipitated PKC zeta-IKK complexes from infected intestinal epithelial cells and recombinant I kappa B alpha as a substrate showed a 2.5-fold increase in I kappa B alpha phosphorylation. PKC zeta can also regulate NF-kappa B by serine phosphorylation of the p65 subunit. Serine phosphorylation of p65 was increased after EPEC infection but could not be consistently attenuated by MYR-PKC zeta-PS, suggesting that other signaling events may be involved in this particular arm of NF-kappa B regulation. We speculate that EPEC infection of intestinal epithelial cells activates several signaling pathways including PKC zeta and ERK that lead to NF-kappa B activation, thus ensuring the proinflammatory response.
Savkovic
Suzana D
SD
Section of Digestive Disease and Nutrition, Department of Medicine, University of Illinois, and West Side Department Of Veterans Affairs Medical Center, Chicago, IL 60612, USA.
Koutsouris
Athanasia
A
Hecht
Gail
G
D018938:Caco-2 Cells; D004751:Enteritis / Q000276:immunology / Q000378:metabolism; D004847:Epithelial Cells / Q000201:enzymology / Q000276:immunology / Q000382:microbiology; D004927:Escherichia coli Infections / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology; D000099221:Protein Kinase C zeta
D020672:I-kappa B Proteins; D020928:Mitogen-Activated Protein Kinases; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B; D011493:Protein Kinase C; D000099221:Protein Kinase C zeta; C083487:I kappa B beta protein; C000606204:NFKBIA protein, human