Autoantibodies reacting against structural proteins of the skin cause severe skin diseases summarized as autoimmune bullous dermatoses (AIBD). One model to study this group of diseases in vivo is the scurfy mouse model. These mice are lacking functional Tregs due to a missense mutation in the forkhead-box-protein 3 (FoxP3) gene. Detailed analyses showed that Scurfy sera contain polyclonal antibodies against several AIBD autoantigens. One of these Scurfy-derived mAb, called 20B12, induces blisters in vivo after injection into newborn mice and binds to the epidermal side of basement membrane zone (BMZ). In further analyses we identified the hemidesmosomal protein BP230 (Bullous Pemphigoid 230 or BPAG1-e), a cytoplasmic protein in basal keratinocytes, as the target antigen of 20B12. First, we used Western Blot analyses to confirm the binding site of 20B12 in the N-terminal domain of BP230. Epitope mapping yielded five putative binding sites within this domain. As 20B12 displays cross-reactivity to human skin and the murine and human BP230 protein show a high degree of homology we further investigated binding of 20B12 to human BP230 protein fragments. Western Blot and ELISA analyses, using fragments covering the five possible epitopes, indicate binding of 20B12 to the Spectrin repeat 9 in the N-terminal domain of BP230. To further elucidate the pathomechanism of blistering we analyzed the involvement of complement system (CS) and Fcγ-Receptors. Preliminary results of 20B12 injection into Fcγ-receptor deficient mice revealed blister formation, indicating Fcγ-receptor-independent mechanism. Indirect immunoflourescene of 20B12 injected WT mice using anti-C3-antibodies yielded no C3 complement deposition at the BMZ suggesting a CS-independent mechanism. In summary, we mapped the binding of the pathogenic murine autoantibody 20B12 to the Spectrin repeat 9 in the N-terminal domain of BP230 and have evidence for a CS- and Fcγ-receptor independent mechanism of blistering.

ESDR012 - Pathogenic murine anti-BP230 autoantibody 20B12: Elucidating the mechanism of blister formation

50th Annual Meeting of the European Society for Dermatological Research. VIRTUAL MEETING