Effect of CD40-P227A polymorphism at human B cell signaling pathways

Objective: To investigate the effects of CD40-P227A SNP at human B cell signaling and function. Methods: Human Ramos B cells were transfected with plasmids of CFP fusion wild type CD40 and mutant type CD40-P227A, together with plasmids of luciferase fusion NF-κB and AP-1, as well as plasmids of GFP fusion NF-κB. Activation of NF-κB and AP-1 pathway were detected by luciferase assay and flow cytometry, degradation of IkBcx and nuclear translocation of NF-κB subunits were tested by Western blot and transfactor ELISA assay. Results: Compared with wild type CD40, CD40-P227A SNP induced more degradation of IκBα, increased nuclear translocation of p65, p50 and c-Rel. As well as higher activity of NF-κB, as shown by increased NF-κB luciferase and GFP-NF-κB expressing B cells. Moreover, CD40-P227A SNP induced more activation of AP-1 pathway than CD40. CD40-P227A enhanced B cells function by inducing more excretion of IL-6 and TNF-α. Conclusion: Our data indicate that CD40-P227A is a gain-of-function phenotype which induces activation of NF-κB and AP-1 signaling pathways and may contribute to the pathogenesis of the SLE autoimmune disease.