Glycogen Synthase Kinase 3α Is the Main Isoform That Regulates the Transcription Factors Nuclear Factor-Kappa B and cAMP Response Element Binding in Bovine Endothelial Cells Infected with .

Glycogen synthase kinase 3 (GSK3) is a constitutive enzyme implicated in the regulation of cytokine expression and the inflammatory response during bacterial infections. Mammals have two GSK3 isoforms named GSK3 alpha and GSK3 beta that plays different but often overlapping functions. Although the role of GSK3 beta in cytokine regulation during the inflammatory response caused by bacteria is well described, GSK3 alpha has not been found to participate in this process. Therefore, we tested if GSK3 alpha may act as a regulatory isoform in the cytokine expression by bovine endothelial cells infected with Staphylococcus aureus because this bacterium is one of the major pathogens that cause tissue damage associated with inflammatory dysfunction. Interestingly, although both isoforms were phosphorylated-inactivated, we consistently observed a higher phosphorylation of GSK3 alpha at Ser21 than that of GSK3 beta at Ser9 after bacterial challenge. During a temporal course of infection, we characterized a molecular switch from pro-inflammatory cytokine expression (IL-8), promoted by nuclear factor-kappa B (NF-kappa B), at an early stage (2 h) to an anti-inflammatory cytokine expression (IL-10), promoted by cAMP response element binding (CREB), at a later stage (6 h). We observed an indirect effect of GSK3 alpha activity on NF-kappa B activation that resulted in a low phosphorylation of CREB at Ser133, a decreased interaction between CREB and the co-activator CREB-binding protein (CBP), and a lower expression level of IL-10. Gene silencing of GSK3 alpha and GSK3 beta with siRNA indicated that GSK3 alpha knockout promoted the interaction between CREB and CBP that, in turn, increased the expression of IL-10, reduced the interaction of NF-kappa B with CBP, and reduced the expression of IL-8. These results indicate that GSK3 alpha functions as the primary isoform that regulates the expression of IL-10 in endothelial cells infected with S. aureus.