Background The B-cell activating factor (BAFF) is a potent survival factor involved in the pathogenesis of autoimmune diseases. Recently, we reported the discovery of a new transcript for BAFF, ∆4BAFF – lacking exon 4 –, which is mainly detected in autoimmune diseases and acts as a transcription factor for its own gene. However, the mechanisms implicated in ∆4BAFF induction and up-regulation are unknown. In this study we analysed the induction and regulation of ∆4BAFF.
Materials and Methods First, to study the alternative splicing of BAFF exon 4, we transfected a minigene construct, centred on exon 4, into RAMOS B cells. To determine the proteins implicated in exon 4 inclusion/exclusion, we co-transfected the minigene together with each of the plasmids coding for the main splicing proteins (SC35, SRp40, SRp55, SRp20 and hnRNPA1), and the ratios between exon4 inclusion/exclusion were evaluated by RT-PCR. Second, we examined the effects of different cytokines on ∆4BAFF induction.
Results RAMOS cells presented exon 4 skipping (ratio inclusion/exclusion: 6.8) after minigene transfection. Following co-transfection of the minigene with coding plasmids for splicing proteins, only the overexpression of SC35 showed effect in the splicing of exon 4, promoting exon 4 inclusion (ratio: >30). Incubation of different cell lines with several cytokines showed that IFN-γ was able to induce ∆4BAFF-transcript. Thus, after IFN-γ stimulation in the minigene model, the ratio inclusion/exclusion markedly decreased (1.5). IFN-γ modifies the balance between SC35 and another member of hnRNPs family (hnRNP C1/C2) favouring the alternative splicing of exon 4.
Conclusions These results demonstrated that IFN-γ induces ∆4BAFF, modifying the function of SC35 protein and increasing the hnRNPC1/C2. Our study provides an expanded conceptual view of BAFF gene regulation, and contributes to a better understanding of the mechanisms involved in BAFF up-regulation in autoimmunity.
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