IL-2 −/− mice develop autoimmunity despite having relatively normal numbers of regulatory T cells (Tregs). In contrast, we demonstrate that IL-2−/− × IL-15−/− and IL-2Rβ−/− mice have a significant decrease in Treg numbers. Ectopic expression of foxp3 in a subset of CD4+ T cells rescued Treg development and prevented autoimmunity in IL-2Rβ−/− mice, suggesting that IL-2Rβ-dependent signals regulate foxp3 expression in Tregs. Subsequent analysis of IL-2Rβ-dependent signal transduction pathways established that the transcription factor STAT5 is necessary and sufficient for Treg development. Specifically, T cell-specific deletion of STAT5 prevented Treg development; conversely, reconstitution of IL-2Rβ−/− mice with bone marrow cells expressing an IL-2Rβ mutant that exclusively activates STAT5 restored Treg development. Finally, STAT5 binds to the promoter of the foxp3 gene suggesting that IL-2Rβ-dependent STAT5 activation promotes Treg differentiation by regulating expression of foxp3.

Appropriate development of regulatory T (Treg) cells is necessary to prevent autoimmunity. Neonatal mice, unlike adults, lack factors required for Treg cell development. It is unclear what these missing factors are. However, signals emanating from the T cell receptor (TCR), the costimulatory receptor CD28, and the family of gammac-dependent cytokine receptors are required for Treg cell development. Herein we demonstrate that expression of a constitutively active Stat5b transgene (Stat5b-CA) allowed for Treg cell development in neonatal mice and restored Treg cell numbers in Cd28(-/-) mice. Sequence analysis of TCR genes in Stat5b-CA Treg cells indicated that ectopic STAT5 activation resulted in a TCR repertoire that more closely resembled that of naive T cells. Using MHCII tetramers to identify antigen-specific T cells, we showed that STAT5 signals diverted thymocytes normally destined to become naive T cells into the Treg cell lineage. Our data support a two-step model of Treg cell differentiation in which TCR and CD28 signals induce cytokine responsiveness and STAT5-inducing cytokines then complete the program of Treg cell differentiation.

Common γ chain (γc)-receptor dependent cytokines are required for regulatory T cell (Treg) development as γc−/− mice lack Tregs. However, it is unclear which γc-dependent cytokines are involved in this process. Furthermore, thymic stromal lymphopoietin (TSLP) has also been suggested to play a role in Treg development. In this study, we demonstrate that developing CD4+Foxp3+ Tregs in the thymus express the IL-2Rβ, IL-4Rα, IL-7Rα, IL-15Rα, and IL-21Rα chains, but not the IL9Rα or TSLPRα chains. Moreover, only IL-2, and to a much lesser degree IL-7 and IL-15, were capable of transducing signals in CD4+Foxp3+ Tregs as determined by monitoring STAT5 phosphorylation. Likewise, IL-2, IL-7, and IL-15, but not TSLP, were capable of inducing the conversion of CD4+CD25+Foxp3− thymic Treg progenitors into CD4+Foxp3+ mature Tregs in vitro. To examine this issue in more detail, we generated IL-2Rβ−/− × IL-7Rα−/− and IL-2Rβ−/− × IL-4Rα−/− mice. We found that IL-2Rβ−/− × IL-7Rα−/− mice were devoid of Tregs thereby recapitulating the phenotype observed in γc−/− mice; in contrast, the phenotype observed in IL-2Rβ−/− × IL-4Rα−/− mice was comparable to that seen in IL-2Rβ−/− mice. Finally, we observed that Tregs from both IL-2−/− and IL-2Rβ−/− mice show elevated expression of IL-7Rα and IL-15Rα chains. Addition of IL-2 to Tregs from IL-2−/− mice led to rapid down-regulation of these receptors. Taken together, our results demonstrate that IL-2 plays the predominant role in Treg development, but that in its absence the IL-7Rα and IL-15Rα chains are up-regulated and allow for IL-7 and IL-15 to partially compensate for loss of IL-2.

Binding of antigen to the B cell antigen receptor (BCR) initiates a multitude of events resulting in B cell activation. How the BCR becomes signaling-competent upon antigen binding is still a matter of controversy. Using a high-resolution proximity ligation assay (PLA) to monitor the conformation of the BCR and its interactions with co-receptors at a 10–20 nm resolution, we provide direct evidence for the opening of BCR dimers during B cell activation. We also show that upon binding Syk opens the receptor by an inside-out signaling mechanism that amplifies BCR signaling. Furthermore, we found that on resting B cells, the coreceptor CD19 is in close proximity with the IgD-BCR and on activated B cells with the IgM-BCR, indicating nanoscale reorganization of receptor clusters during B cell activation.DOI: http://dx.doi.org/10.7554/eLife.02069.001