Evidence for AIRE Binding to Promoter Regions of Known Autoantigens in Human Peripheral Blood by Chromatin Immunoprecipitation Assay

Central tolerance is provided by the AIRE-expressing medullary thymic epithelial cells, through high avidity recognition of self-antigens. Nevertheless, peripheral mechanisms regulate adaptive immunity by deleting autoreactive T-cells that escape thymic selection or inducing their functional unresponsiveness. These mechanisms require interaction with antigen presenting cells exposing cognate antigen. As regard multiple types of extrathymic AIRE-expressing cells, residing in secondary lymphoid organs, were described. In this study we aimed to provide evidence for AIRE binding to promoter regions of known autoantigens in human peripheral blood mononuclear cells (PBMC) in an attempt to elucidate whether this non-classical transcriptional factor could play a role in the pe-ripheral expression of self-antigens. Chromatin immunoprecipitation (ChIP) of 4 normal human PBMC samples was performed using anti-AIRE monoclonal antibody. Quantitative-Real-Time PCR (qRT-PCR) was used to detect AIRE-binding at promoters of known autoantigens, including thyroidrelated thyroglobulin, thyroperoxidase, thyrotropin-receptor, Type 1 diabetes-related autoantigens, i.e. insulin and zinc transporter 8, and to confirm their expression in PBMC. ChIP evidenced amplicons of promoter regions of mentioned autoantigens by qRT-PCR. Expression of AIRE and of autoantigens was confirmed in the same human PBMC samples. This study provides the first evidence that AIRE binds promoters of known autoantigens in human PBMC, supporting its expression and potential role in modulating peripheral self-antigen expression.