The central helicase domain holds the major conformational epitopes of melanoma differentiation–associated gene 5 autoantibodies

Abstract

Objectives Autoantibodies against MDA5 serve as a biomarker for dermatomyositis (DM) and a risk factor for interstitial lung disease (ILD). MDA5 is a protein responsible for sensing RNA virus infection and activating signalling pathways against it. However, little is known about antigen epitopes on MDA5 autoantibodies. We aimed to determine the interaction of the MDA5 autoantibody-antigen epitope. Methods Cell-based assays (CBAs), immunoprecipitation-immunoblot assays, and various immunoblotting techniques were used in the study. Results We demonstrate that DM patient autoantibodies recognize MDA5 epitopes in a native conformation-dependent manner. Furthermore, we identified the central helicase domain formed by Hel1, Hel2i, Hel2, and pincer (3Hel) as the major epitopes. As proof of principle, the purified 3Hel efficiently absorbed MDA5 autoantibodies from patient sera through immunoprecipitation-immunoblot assay. Conclusion Our study uncovers the nature of antigen epitopes on MDA5 and provides guidance for diagnosis and targeted therapeutic approach development.