Effect of the Aire gene on the chemotactic property of medullary thymic epithelial cells on thymocytes

Abstract

The Autoimmune regulator (Aire) gene (human chromosome 21q22.3 and Mus musculus 10qC1), is a controller of central immune tolerance and the emergence of autoimmune diseases. The Aire sequence presents ~ 80% man-mouse similarity and this gene regulates the transcription of peripheral tissue autoantigen genes (PTAs) in medullary thymic epithelial cells (mTECs). This phenomenon was termed promiscuous gene expression (PGE). The meaning of PGE is immunological, that is, the self-representation in the thymus, which is the basis of the induction of immunological tolerance. For autoantigen tolerance to occur, thymocytes (the precursors of T cells) originating in the bone marrow migrate to the thymus and enter this organ through the cortico-medullary junction. Once within the thymus, the thymocytes migrate sequentially and in that order, cortex and then medulla, establishing respectively adhesion with cortical thymic epithelial cells (cTEC-thymocyte) and then with mTECs (mTEC-thymocyte). These interactions are commonly referred to as thymic crosstalk. These migration and adhesion events mediate the positive selection (cTECs) and negative selection (mTECs) of thymocytes. The migration of thymocytes into the thymus is mediated chemokines such as CXCL12 (secreted by cTECs) and CCL19 and CCL21 (secreted by mTECs). In our laboratory we are investigating the role of the Aire gene in mTEC cells through the loss of function approach, inducing deleterious mutations by the Crispr-Cas9 system. The Aire mutant cells, deregulate a large set of downstream genes including those encoding adhesion molecules. In this project we intend to investigate whether Aire also controls chemokine genes involved in the migration of thymocytes toward mTEC cells. For this, we use mTEC cells in culture, comparing the transcriptional expression of wild-type mTECs vs Aire mutant mTECs. The functional validation would be performed by the thymocyte - mTEC migration (transwell assay).