Dynamic transcriptional activity and chromatin remodeling of regulatory T cells after varied duration of IL-2R signaling

Regulatory T cells (Tregs) are necessary for the maintenance of immune self-tolerance and homeostasis. They have a heightened sensitivity to IL-2, which may create a therapeutic window to promote immune regulation by their selective stimulation. While IL-2 responsive genes are well characterized, yet remain unknown genetic, chromatin and metabolic programs during sustained IL-2R signaling. Longitudinal transcriptional and chromatin accessibility profiling and metabolic studies were performed on peripheral Tregs in response to long-lived mIL-2/CD25 fusion protein. We found an initially increased STAT5 dependent gene modulation enabled by enhanced chromatin accessibility, whereas a STAT5-independent genome-wide chromatin closing to inhibit transcription and cell signaling occurred later. Interestingly, IL-2-dependent genes were induced upon re-stimulation. Mitochondrial metabolism was reprogrammed to fulfill the bioenergetics demand for the burst of proliferation. We propose that a shift from euchromatin to heterochromatin represents a mechanism to restrain, but not abrogate, persistent IL-2R signaling to maintain Treg homeostasis. Overall design: Mouse spleen Tregs DNA accessibility of mIL-2, mIL-2/CD25 (FP) or control PBS injected mice were generated 0.75 hr post injection by deep sequencing in triplicate using NextSeq 500 with a High Output Kit 150-cycle flow cell (Illumina). Reads from ATAC-seq were mapped to the Mus musculus genome GRCm38 using bwa-mem and MACS was used to identify the peak regions. The Irreproducible Discovery Rate (IDR) method was used to identify reproducible peaks between two pseudo-replicates that were generated for each sample. Only reproducible peaks (IDR > 0.1) were retained for downstream analyses. Differential expressed genes in Tregs between the mIL-2 or mIL-2/CD25 and PBS injected mice were identified using DESeq2, and determined according to threshold of p <0.05