Regulatory T cells suppress the formation of super-effector CD8 T cells by limiting IL-2

In this experiment, the gene expression of CD8+ T cells primed in the presence or absence of Tregs on a single cell level was analyzed. For this purpose, Ly5.1 OT-I T cells were adoptively transferred into Treg deficient (DEREG+) or Treg replete (DEREG-) mice, activated with DC-OVA, isolated after 3 days, and analyzed by scRNAseq. Invididual samples were indexed with oligo-tagged TotalSeq-C antibodies and pooled prior to the analysis. We observed that Tregs reduced the expression of IL-2 responsive genes, including key cytotoxic molecule granzyme B. Unsupervised clustering revealed 5 clusters including a cluster of cells which did not match any usual CD8+ subsets and, due to unusual co-expression of effector molecules such as GZMK and KLRK1, were dubbed super-effector cells. Overall design: Ly5.1 OT-I T cells were adoptively transferred into double-transgenic Treg depleted DEREG+ RIP.OVA and Treg control DEREG- RIP.OVA mice (C57BL6 background, n=3 per group). The following day the mice were immunized with DC-OVA. Third day after the immunization their spleens were isolated, the OT-I cells were FACS sorted as Ly5.1+ CD8+ cells, and analyzed via scRNA-seq using 10X Genomics Single Cell Immune Profiling with Feature Barcoding.