Negative regulation of Lyn protein-tyrosine kinase by c-Cbl ubiquitin-protein ligase in FcεRI-mediated mast cell activation

Background: Recent studies have demonstrated that c-Cbl functions as a ubiquitin-protein ligase toward immune receptors and non-receptor protein-tyrosine kinase Syk by facilitating their ubiquitination and subsequent targeting to proteasomes. However, it was not clear whether Src family kinase Lyn is regulated by the Cbl family of ubiquitin-protein ligases.<LF> <LF>Results: Aggregation of the high affinity IgE receptor (FcepsilonRI) induces the rapid ubiquitination of Lyn in rat basophilic leukaemia RBL-2H3 cells. Treatment of cells with a proteasome inhibitor enhances the ubiquitination of Lyn. Stimulation of FcepsilonRI results in the association of Lyn with c-Cbl and Cbl-b, both of which then become tyrosine phosphorylated. Co-transfection study shows that both c-Cbl and Cbl-b could induce the ubiquitination of activated Lyn in COS cells. Furthermore, over-expression of membrane-anchored form of c-Cbl inhibits the FcepsilonRI-mediated degranulation and cytokine gene production in RBL-2H3 cells by the down-regulation of the kinase activity of Lyn through the enhanced ubiquitination. Conclusions: These results demonstrate that Lyn is down-regulated by c-Cbl-mediated ubiquitination and subsequent degradation in proteasome after FcepsilonRI stimulation in mast cells. Targeting of c-Cbl in the lipid raft results in the inhibition of FcepsilonRI-mediated mast cell activation.