Molecular Mechanisms Underlying FOXP3 Induction in Human T Cells

Abstract

FOXP3 is playing an essential role for T regulatory cells and is involved in the molecular mechanisms controlling immune tolerance. Although the biological relevance of this transcription factor is well documented, the pathways responsible for its induction are still unclear. The current study reveals structure and function of the human FOXP3 promoter, revealing essential molecular mechanisms of its induction. The FOXP3 promoter was defined by RACE, cloned, and functionally analyzed using reporter-gene constructs in primary human T cells. The analysis revealed the basal, T cell-specific promoter with a TATA and CAAT box 6000 bp upstream the translation start site. The basal promoter contains six NF-AT and AP-1 binding sites, which are positively regulating the trans activation of the FOXP3 promoter after triggering of the TCR. The chromatin region containing the FOXP3 promoter was bound by NF-ATc2 under these conditions. Furthermore, FOXP3 expression was observed following TCR engagement. Promoter activity, mRNA, and protein expression of T cells were suppressed by addition of cyclosporin A. Taken together, this study reveals the structure of the human FOXP3 promoter and provides new insights in mechanisms of addressing T regulatory cell-inducing signals useful for promoting immune tolerance. Furthermore, the study identifies essential, positive regulators of the FOXP3 gene and highlights cyclosporin A as an inhibitor of FOXP3 expression contrasting other immunosuppressants such as steroids or rapamycin.

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

“…It is also important to note that our results do not dispute the crucial role of TCR in human Treg cell commitment. Analysis of the FOXP3 promoter area has identified several TCR-responsive sites (26) and TCR ligation can lead to up-regulation of FOXP3 (27), although this is not necessarily accompanied by suppressive function (28 -30). In all likelihood the posited Treg cell maturation pathway involves TCR-dependent checkpoints, perhaps selecting for autoreactive clones among a stochastically generated FOXP3 ϩ precursor pool (3-7).…”

FOXP3+ regulatory T cells: Current controversies and future perspectives

“…In contrast, several studies have reported that FOXP3 mRNA is expressed by in vitro human activated non-regulatory T cells [14,[20][21][22], which may gain regulatory activity [20,22]. Recent studies on the FOXP3 promoter identify a non-repressive chromatin structure that together with a specific transcriptional regulation programme may explain the ability of activation to induce FOXP3 expression in CD4 + CD25 -T cells [23]. Gavin and colleagues [24] have recently reported that activation-induced FOXP3 expression in human T cells, as measured by flow cytometry, is transient and insufficient to induce Treg, proposing that sustained high level FOXP3 expression is required for Treg phenotype and function.…”

Rapamycin, not cyclosporine, permits thymic generation and peripheral preservation of CD4+CD25+FoxP3+ T cells

“…11,28 CsA can inhibit this thymic generation of CD4 þ FoxP3 þ T REG by impairment of TCR signaling, and more specifically by reducing the NFATdependent FoxP3 promotor activity. 29 Rapamycin inhibits biochemical pathways required for the cell cycle progression from the G1 to S phase, but allows the initial signal transduction upon TCR triggering. 15 We found that rapamycin does not affect the thymic CD4 þ FoxP3 þ T-cell generation.…”