The Autoimmune Regulator (Aire) transactivates HLA-G gene expression in thymic epithelial cells

The Autoimmune Regulator (Aire) protein coordinates the negative selection of developing thymocytes by inducing the expression of hundreds of tissue-specific antigens within the thymic medulla, which is also a primary site of the expression of the immune checkpoint HLA-G molecule. Considering the immunomodulatory properties of Aire and HLA-G, and considering that the role of the constitutive thymus expression of HLA-G has not been elucidated, we studied the effect of AIRE cDNA transfection on HLA-G expression in 4D6 thymic cells and in the HLA-G-positive JEG-3 choriocarcinoma cells. Aire promoted the transactivation of HLA-G gene by increasing the overall transcription, inducing the transcription of at least G1 and G2/G4 isoforms, and incrementing the occurrence and distribution of intracellular HLA-G protein solely in 4D6 thymic cells. Luciferase-based assays and chromatin immunoprecipitation experiments performed in 4D6 cells revealed that Aire targeted at least two regions within the 5'-untranslated regulatory region (5'-URR) extending 1·4 kb from the first ATG initiation codon. The interaction occurs independently of three putative Aire-binding sites. These results indicate that the Aire-induced upregulation of HLA-G in thymic cells is likely to act through the interaction of Aire with specific HLA-G 5'-URR DNA-binding factors. Such a multimeric transcriptional complex might operate in the thymus during the process of promiscuous gene expression.