Foxp3-dependent programme of regulatory T-cell differentiation

Abstract

Regulatory CD4 1 T cells (T R cells), the development of which is critically dependent on X-linked transcription factor Foxp3 (forkhead box P3), prevent self-destructive immune responses 1 . Despite its important role, molecular and functional features conferred by Foxp3 to T R precursor cells remain unknown. It has been suggested that Foxp3 expression is required for both survival of T R precursors as well as their inability to produce interleukin (IL)-2 and independently proliferate after T-cell-receptor engagement, raising the possibility that such 'anergy' and T R suppressive capacity are intimately linked 2-4 . Here we show, by dissociating Foxp3-dependent features from those induced by the signals preceding and promoting its expression in mice, that the latter signals include several functional and transcriptional hallmarks of T R cells. Although its function is required for T R cell suppressor activity, Foxp3 to a large extent amplifies and fixes pre-established molecular features of T R cells, including anergy and dependence on paracrine IL-2. Furthermore, Foxp3 solidifies T R cell lineage stability through modification of cell surface and signalling molecules, resulting in adaptation to the signals required to induce and maintain T R cells. This adaptation includes Foxp3-dependent repression of cyclic nucleotide phosphodiesterase 3B, affecting genes responsible for T R cell homeostasis.In males, Foxp3 deficiency results in fatal early-onset systemic autoimmune disease 5 . In heterozygote Foxp3 wt/null females only one-half of T cells harbours the mutant Foxp3 allele due to random X-chromosome inactivation, whereas autoimmunity is controlled by a normal T R population expressing the Foxp3 wild-type allele. Thus, we were able to genetically mark cells actively transcribing a Foxp3 null allele, yet lacking Foxp3 protein (hereafter called T FN for Foxp3 nullexpressing T cells), through an in-frame insertion of GFP into a stop-codon-disrupted Foxp3 locus (Foxp3 gfpko ) and investigate their features in mice ( Fig. 1a; see also Supplementary Figs 1 and 2a). Female Foxp3 gfpko/wt mice were healthy, whereas male Foxp3 gfpko mice developed the same severity of autoimmunity as Foxp3 knockout (Foxp3 null ) mice 6 , resulting in death at ,4 weeks of age. Thymocyte and peripheral lymphoid organ cellularity did not differ between Foxp3 gfpko/wt and Foxp3 gfp/gfp mice, nor did the proportion of Foxp3 1 T R cells and Foxp3 2 CD4 1 T cells (data not shown). As our main focus was to characterize T FN cells in healthy Foxp3 gfpko/wt mice, analysis of autoimmune male Foxp3 gfpko mice is included as Supplementary Fig. 2.T FN cells constituted ,1-3% of mature CD4 1 thymocytes and peripheral CD4 1 T cells, indicating that Foxp3 is not required to rescue T R precursors from negative selection (Fig. 1b, c). This is consistent with a reported abundance of T-cell receptors (TCRs) characteristic of T R cells in Foxp3 null mice 7 . As ectopic expression of Foxp3 has been shown to induce a state of hyporesponsiveness in CD4 1 T cell...

Cutting Edge: Human CD4−CD8− Thymocytes Express FOXP3 in the Absence of a TCR

“…Although our results are at variance with murine data, recent work suggests that in mice also the induction of a Treg cell genetic program may be an early event (23) and precedes the expression of FoxP3 (24,25). It is also important to note that our results do not dispute the crucial role of TCR in human Treg cell commitment.…”

Agonist-Driven Development of CD4+CD25+Foxp3+ Regulatory T Cells Requires a Second Signal Mediated by Stat6

“…The IL-4/STAT6/c-Maf/CD25 and IL-2/ CD25/c-Maf/IL-4 pathways may converge to act on CD25 ϩ Foxp3 ϩ Treg cells, but their relative roles remain to be established. It was recently shown that STAT5 is required for Foxp3 induction (18,19) and Foxp3 may be at the crossroads to Th2 or Th17 differentiation (33,34). In the thymus, Ag-specific Treg cell development occurs in the absence of IL-2 (24), indicating that the STAT6 pathway may act independently of IL-2 and that TCR and STAT6 signals suffice for efficient Treg cell generation.…”

Is FOXP3 a bona fide marker for human regulatory T cells?

“…That FOXP3 is not only a marker of murine nTreg but that it is also necessary for their function is a broadly accepted concept. This is due to experimental evidence indicating that constitutive and specific FOXP3 expression, which cannot be induced upon TCR stimulation [2], is essential for stabilizing and amplifying regulatory functions that include the induction of anergy and the impairment of IL-2 production [8]. Conversely, the notion that FOXP3 expression can be detected in human T cells other than nTreg has recently questioned the role of this molecule as a master regulator of nTreg biology in man.…”