I n the periphery, F oxp3 expression is considered sufficient to maintain natural regulatory CD 4 + T ‐cell suppressive function. In this study, we challenge this model. Indeed, in mouse chimeras in which major histocompatibility complex ( MHC ) class II expression is restricted to the thymus, peripheral regulatory CD 4 + T cells lack suppressive activity. In addition, regulatory CD 4 + T cells recovered 5 days after transfer into recipient mice lacking expression of MHC class II molecules (self‐deprived) are unable to inhibit the proliferative response of conventional CD4 + T cells both in vitro and in vivo. Disruption of TCR / MHC class II interactions rapidly leads to alterations in the regulatory CD 4 + T ‐cell phenotype, the ability to respond to stimulation and to produce interleukin‐10, and the transcriptional signature. Interestingly, self‐deprivation does not affect Foxp3 expression indicating that in regulatory CD4 + T cells, self‐recognition induces unique transcriptional and functional features that do not rely on Foxp3 expression.