miR-155 exerts posttranscriptional control of autoimmune regulator (Aire) and tissue-restricted antigen genes in medullary thymic epithelial cells

Background: The autoimmune regulator (Aire) gene is critical for the appropriate establishment of central immune tolerance. As one of the main controllers of promiscuous gene expression in the thymus, Aire promotes the expression of thousands of downstream tissue-restricted antigen (TRA) genes, cell adhesion genes and transcription factor genes in medullary thymic epithelial cells (mTECs). Despite the increasing knowledge about the role of Aire as an upstream transcriptional controller, little is known about the mechanisms by which this gene could be regulated. Results: Here, we assessed the posttranscriptional control of Aire by miRNAs. The in silico miRNA-mRNA interaction analysis predicted thermodynamically stable hybridization between the 3UTR of Aire mRNA and miR-155, which was confirmed to occur within the cellular milieu through a luciferase reporter assay. This finding enabled us to hypothesize that miR-155 might play a role as an intracellular posttranscriptional regulator of Aire mRNA. To test this hypothesis, we transfected a murine mTEC cell line with a miR-155 mimic in vitro, which reduced the mRNA and protein levels of Aire. Moreover, large-scale transcriptome analysis showed the modulation of 311 downstream mRNAs, which included 58 TRA mRNAs. Moreover, miR-155 mimic-transfected cells exhibited a decrease in their chemotaxis property compared with control thymocytes. Conclusion: Overall, the results indicate that miR-155 may posttranscriptionally control Aire mRNA as well as a crucial process by which mTECs allow migration of thymocytes through chemotaxis.

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Autoimmune regulator act in synergism with thymocyte adhesion in the control of lncRNAs in medullary thymic epithelial cells

10.1101/2021.07.14.452339 ◽

2021 ◽

Author(s):

Max J Duarte ◽

Romario S Mascarenhas ◽

Amanda F Assis ◽

Pedro P Tanaka ◽

Cesar A Speck-Hernandez ◽

...

Keyword(s):

Epithelial Cells ◽

Model Systems ◽

Thymic Epithelial Cells ◽

Large Set ◽

Rna Pol Ii ◽

Pol Ii ◽

Autoimmune Regulator ◽

Medullary Thymic Epithelial Cells

The autoimmune regulator (Aire) gene in medullary thymic epithelial cells (mTECs) encodes the AIRE protein, which interacts with its partners within the nucleus. This Aire complex induces stalled RNA Pol II on chromatin to proceed with transcription elongation of a large set of messenger RNAs and microRNAs. Considering that RNA Pol II also transcribes long noncoding RNAs (lncRNAs), we hypothesized that Aire might be implicated in the upstream control of this RNA species. To test this, we employed a loss-of-function approach in which Aire knockout mTECs were compared to Aire wild-type mTECs for lncRNA transcriptional profiling both in vitro and in vivo model systems. RNA sequencing enables the differential expression profiling of lncRNAs when these cells adhere in vitro to thymocytes or do not adhere to them as a way to test the effect of cell adhesion. Sets of lncRNAs that are unique and that are shared in vitro and in vivo were identified. Among these, we found the Aire-dependent lncRNAs as for example, Platr28, Ifi30, Morrbid, Malat1, and Xist. This finding represents the first evidence that Aire mediates the transcription of lncRNAs in mTECs. Microarray hybridizations enabled us to observe that temporal thymocyte adhesion modulates the expression levels of such lncRNAs as Morrbid, Xist, and Fbxl12o after 36h of adhesion. This finding shows the existence of a synergistic mechanism involving a link between thymocyte adhesion, Aire, and lncRNAs in mTECs that might be important for immune self-representation.

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