Characterization of long non-coding RNA controlling regulatory T cell identity
ADOUE V. 1, ZAHM M. 1, LAVIOLETTE K. 1, GHAZALI S. 1, LEBEURRIER M. 1, JOFFRE O. 1
1 Infinity - Toulouse Institute For Infectious and Inflammatory Diseases INSERM UMR1291 – CNRS UMR5051 – Université Toulouse III, Toulouse, France
Conventional CD4+ T lymphocytes (Tconv) play a central role in protecting the host against a wide range of dangers. Their activity needs however to be tightly controlled by a subpopulation of CD4+ regulatory T cells (Treg) endowed with immunosuppressive functions and expressing the transcription factor Foxp3. However, while necessary, Foxp3 is not sufficient to define Treg cell identity. LncRNAs are cell-type specific regulators that critically shape transcriptional networks. However, the current lncRNAs annotation is mostly based on cell lines or whole organisms and the Treg-specific lncRNA repertoire largely remains to be characterized. Using a transgenic mouse model, we isolated 5 Treg subpopulations and their Tconv counterparts based on location, origin and activation status. Using deep RNA sequencing and refined bioinformatic pipelines, we identified >2000 new lncRNA’s genes not previously annotated in databases. We segregated the lncRNAs into three modules based on their expression across the 10 T-cell subpopulations and identified the key transcription factors controlling their expression. Importantly, we identified a Treg-identity module (“Treg-Id”), which encompasses 49 lncRNAs specifically expressed in Tregs, located at Treg-specific super-enhancers, and whose expression is controlled by the chromatin modifier Satb1. We showed that Treg-Id’s lncRNAs are associated with enhancer-promoter loops at Treg signature protein-coding genes. We finally used CRISPR/Cas9 to delete “Lcincr”, a previously unknown lncRNA located in the same locus than GARP, and showed that its deletion is associated with an upregulation of GARP (+300%) and Foxp3 expression. Overall, we characterized the lncRNA landscape in Treg subpopulations and identified Lcincr, a new lncRNA controlling GARP expression.