T cell receptor (TCR) signals can elicit full activation with acquisition of effector functions or a state of anergy. Here, we ask whether microRNAs affect the interpretation of TCR signaling. We find that Dicer-deficient CD4 T cells fail to correctly discriminate between activating and anergy-inducing stimuli and produce IL-2 in the absence of co-stimulation. Excess IL-2 production by Dicer-deficient CD4 T cells was sufficient to override anergy induction in WT T cells and to restore inducible Foxp3 expression in Il2-deficient CD4 T cells. Phosphorylation of Akt on S473 and of S6 ribosomal protein was increased and sustained in Dicer-deficient CD4 T cells, indicating elevated mTOR activity. The mTOR components Mtor and Rictor were posttranscriptionally deregulated, and the microRNAs Let-7 and miR-16 targeted the Mtor and Rictor mRNAs. Remarkably, returning Mtor and Rictor to normal levels by deleting one allele of Mtor and one allele of Rictor was sufficient to reduce Akt S473 phosphorylation and to reduce co-stimulation–independent IL-2 production in Dicer-deficient CD4 T cells. These results show that microRNAs regulate the expression of mTOR components in T cells, and that this regulation is critical for the modulation of mTOR activity. Hence, microRNAs contribute to the discrimination between T cell activation and anergy.

Runx proteins are essential for hematopoiesis and play an important role in T cell development by regulating key target genes, such as CD4 and CD8 as well as lymphokine genes, during the specialization of naive CD4 T cells into distinct T helper subsets. In regulatory T (T reg) cells, the signature transcription factor Foxp3 interacts with and modulates the function of several other DNA binding proteins, including Runx family members, at the protein level. We show that Runx proteins also regulate the initiation and the maintenance of Foxp3 gene expression in CD4 T cells. Full-length Runx promoted the de novo expression of Foxp3 during inducible T reg cell differentiation, whereas the isolated dominant-negative Runt DNA binding domain antagonized de novo Foxp3 expression. Foxp3 expression in natural T reg cells remained dependent on Runx proteins and correlated with the binding of Runx/core-binding factor β to regulatory elements within the Foxp3 locus. Our data show that Runx and Foxp3 are components of a feed-forward loop in which Runx proteins contribute to the expression of Foxp3 and cooperate with Foxp3 proteins to regulate the expression of downstream target genes.

Here, we report data concerning the discovery in adult human peripheral blood of a precursor cell population able to differentiate into CD4⁺CD3⁺αβ⁺ mature T cells. These cells, which represent 0.1–0.5% of total peripheral blood mononuclear cells (PBMC), express substantial levels of CD4, but lack CD3 surface expression. At a molecular level, they express the pre-T cell receptor α (pTα) gene, CD3-γ, CD-δ and CD-ε, and RAG-1 recombination enzyme and have initiated rearrangements in the T cell receptor (TCR)-β locus (D–J). Moreover, low levels of CD3ε protein, but not of TCR-β chain, can be detected in their cytoplasm. Our results suggest that CD4⁺CD3⁻ cells identified in peripheral blood are different from CD3⁻CD4⁺CD8⁻ thymocytes and may contain precursors of an extrathymic T cell differentiation pathway.