For Day 0 Tregs, spleens of Foxp3(IRES-mRFP) mice were harvested and stained with CD4 antibody to sort CD4+ RFP+ Tregs. For tthe Day 3 recovered exogenous Tregs, C57BL6/J mice with tissue injuries (in bone, muscle, skin and heart tissues) that were treated with RFP+ Tregs from Foxp3(IRES-mRFP) mice were harvested, and subjected to enzymatic tissue digestion, followed by subsequent staining with surface antibodies for cell sorting. The recovered Tregs were purified by first excluding any dead cells, then excluding any potential contaminating CD11b, F4/80, Ly6G-positive myeloid cells, followed by selecting CD3, CD4 and RFP+ Tregs. All cells were directly sorted into the primer/lysis buffer mix in 384-well plates (50 cells/well) and then frozen for further processing. Cells from the same sample that were sorted in multiple wells (in increments of 50 cells per well) were later combined bioinformatically to create pseudobulked samples. Minibulk transcriptome libraries were generated using a modified CEL-Seq2 protocol (Hashimshony et al., 2016, Genome Biol.) with the following adaptations: Samples were pooled after first strand cDNA synthesis, treated with Exonuclease 1 for 30 minutes, followed by a 1.2X bead clean-up. Second strand synthesis was performed using NEBNext Second Strand Synthesis module (NEB #E6111S) in a final reaction volume of 20 ul and NucleoMag NGS Clean-up and Size select magnetic beads (Macherey-Nagel - 7449970.5) were used for all DNA purification and size selection steps.