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...

SUMMARY Resveratrol, a polyphenol in red wine, has been reported as a calorie restriction mimetic with potential antiaging and antidiabetogenic properties. It is widely consumed as a nutritional supplement, but its mechanism of action remains a mystery. Here, we report that the metabolic effects of resveratrol result from competitive inhibition of cAMP-degrading phosphodiesterases, leading to elevated cAMP levels. The resulting activation of Epac1, a cAMP effector protein, increases intracellular Ca2+ levels and activates the CamKKβ-AMPK pathway via phospholipase C and the ryanodine receptor Ca2+-release channel. As a consequence, resveratrol increases NAD+ and the activity of Sirt1. Inhibiting PDE4 with rolipram reproduces all of the metabolic benefits of resveratrol, including prevention of diet-induced obesity and an increase in mitochondrial function, physical stamina, and glucose tolerance in mice. Therefore, administration of PDE4 inhibitors may also protect against and ameliorate the symptoms of metabolic diseases associated with aging.

Cyclic nucleotide phosphodiesterases (PDEs) catalyse the hydrolysis of cyclic AMP and cyclic GMP, thereby regulating the intracellular concentrations of these cyclic nucleotides, their signalling pathways and, consequently, myriad biological responses in health and disease. Currently, a small number of PDE inhibitors are used clinically for treating the pathophysiological dysregulation of cyclic nucleotide signalling in several disorders, including erectile dysfunction, pulmonary hypertension, acute refractory cardiac failure, intermittent claudication and chronic obstructive pulmonary disease. However, pharmaceutical interest in PDEs has been reignited by the increasing understanding of the roles of individual PDEs in regulating the subcellular compartmentalization of specific cyclic nucleotide signalling pathways, by the structure-based design of novel specific inhibitors and by the development of more sophisticated strategies to target individual PDE variants.

cAMP and cGMP mediate biological responses initiated by diverse extracellular signals. By catalyzing hydrolysis of the 3Ј-5Ј-phosphodiester bond of cyclic nucleotides, cyclic nucleotide phosphodiesterases (PDEs) 1 regulate intracellular concentrations and effects of these second messengers. PDEs include a large group of structurally related enzymes (reviewed in Refs.