Nuclear factor of activated T cells (NFAT) is implicated in multiple biologicalNuclear factor of activated T cells (NFAT) is a group of transcription factors that was first identified to play an important role in cytokine gene expression (22). Subsequent studies demonstrated that NFATs are present in numerous tissues (33,34,40). The wide tissue distribution of the NFAT isoforms suggests that NFAT may participate in multiple physiological processes. Recently, NFAT activity has been implicated in adipocyte differentiation, cardiac hypertrophy, and learning and memory (30,32,41). Thus, elucidation of mechanisms that regulate NFAT is critical for understanding these biological processes.Four distinct genes encoding closely related NFAT proteins (NFATc1/NFATc/NFAT2, NFATc2/NFATp/NFAT1, NFATc3/ NFAT4/NFATx, and NFATc4/NFAT3) have been identified (reviewed in references 18 and 47). Alternative mRNA splicing of these four genes further generates at least 10 different NFAT polypeptides. The function of these alternatively spliced NFAT isoforms remains elusive. However, all NFAT members contain a highly conserved NH 2 -terminal regulatory NFAT homology domain and a COOH-terminal Rel homology region for DNA binding. Thus, understanding the function of these conserved domains will provide new insights on NFAT regulation.The NH 2 -terminal NFAT homology domain encodes several distinct sequences, including the PXIXIT motif, the Ser-rich region (SRR), and the Ser-Pro (SP)-rich boxes for NFAT regulation (18, 47). These sequences are found in all NFAT members. The PXIXIT motif is recognized by the calcineurin phosphatase (3, 15), which dephosphorylates NFAT upon activation. Sequestration of the calcineurin phosphatase by overexpression of the PXIXIT motif blocks NFAT activation. The SRR and the SP boxes are major targets for NFAT phosphorylation (4-6, 12-14, 44, 46, 58). Dephosphorylation of Ser residues in the SRR and the SP boxes promotes nuclear localization of NFAT. Thus, dephosphorylation of the NFAT homology domain, which is mediated by the calcineurin phosphatase, plays an important role in NFAT activation.Once NFAT is dephosphorylated and translocated into the nucleus, activated NFAT interacts with other transcription factors to induce gene expression. The interaction of NFAT with Fos-Jun (AP-1 complex), GATA, and MEF2 suggests that NFAT often functions at composite DNA elements (7,41,43,54,56). Formation of a ternary complex induces expression of NFAT targets, such as interleukin-2 (IL-2), IL-4, IL-5, and tumor necrosis factor alpha. However, physiological function of NFAT in nonimmune tissues remains to be established.Multiple protein kinases, including the mitogen-activated protein (MAP) kinase group (ERK, JNK, and p38 kinase), glycogen synthase kinase 3␤ (GSK3␤), protein kinase A (PKA), and casein kinase 1␣ (CK1␣), have been shown to phosphorylate NFAT (4,6,[12][13][14]29,46,58). NFAT is phosphorylated on multiple Ser residues located in the conserved SRR and the SP boxes. Phosphorylation of these Ser residues oppose...

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ADP-ribosylation is a reversible posttranslational modification mediated by poly-ADP-ribose polymerase (PARP). The results of recent studies demonstrate that ADP-ribosylation contributes to transcription regulation. Here, we report that transcription factor NFAT binds to and is ADP-ribosylated by PARP-1 in an activation-dependent manner. Mechanistically, ADP-ribosylation increases NFAT DNA binding. Functionally, NFAT-mediated interleukin-2 (IL-2) expression was reduced in T cells upon genetic ablation or pharmacological inhibition of PARP-1. Parp-1 ؊/؊ T cells also exhibit reduced expression of other NFAT-dependent cytokines, such as IL-4. Together, these results demonstrate that ADP-ribosylation mediated by PARP-1 provides a molecular switch to positively regulate NFAT-dependent cytokine gene transcription. These results also imply that, similar to the effect of calcineurin inhibition, PARP-1 inhibition may be beneficial in modulating immune functions.ADP-ribosylation is a reversible posttranslational modification that transfers ADP-ribose from NAD ϩ to Glu, Asp, and/or Arg amino acids of target proteins (18). Similar to ubiquitination, ADP-ribosylation modifies target proteins to various masses due to the assorted chain lengths of the ADPribose. ADP-ribosylation is inhibited by the NAD ϩ analog 3-aminobenzamide and, more specifically, by PJ-34 (45). Poly-ADP-ribose polymerase-1 (PARP-1) is a nuclear enzyme that accounts for the bulk of ADP-ribosylation in vivo (43). Indeed, only ϳ10% of PARP activity remains in Parp-1 Ϫ/Ϫ cells upon DNA damage. In addition to its role in DNA damage repair, the results of recent studies demonstrate that PARP-1 contributes to gene transcription regulation (26,40).Transcription factor NFAT is the master regulator of interleukin-2 (IL-2) gene transcription (24,42). In resting cells, NFAT resides in the cytosol. The nuclear accumulation of NFAT is regulated by calcineurin-mediated dephosphorylation (9, 15, 23). The immunosuppressant drugs cyclosporine A (CsA) and tacrolimus (FK506) inhibit calcineurin and abrogate NFAT activation. Indeed, understanding the mechanism of NFAT activation has contributed to the great advances in transplantation surgery (27). Given that immunosuppressant therapy using CsA or FK506 causes neuro-and nephrotoxicity (1, 19), further understanding of the molecular basis of NFAT activation will provide alternate therapeutic targets for the treatment of transplant patients.Once in the nucleus, NFAT interacts with coregulators to achieve optimal NFAT activation (11,21,28). These NFAT coregulators include Fos-Jun, C/EBPs, and Fox3p, which form a composite transcription complex to regulate NFAT-mediated gene transcription. In addition, transcription coactivator CREBbinding protein/p300 and class II histone deacetylases are recruited to modulate NFAT-mediated transcription (3,12,16,48). Here, we report that PARP-1 binds to and ADP-ribosylates NFAT. The ADP-ribosylation mediated by PARP-1 provides a molecular switch to positively regulate NFAT-dependent cytokine ge...

Compromised immunoregulation contributes to obesity and complications in metabolic pathogenesis. Here, we demonstrate that the nuclear factor of activated T cell (NFAT) group of transcription factors contributes to glucose and insulin homeostasis. Expression of two members of the NFAT family (NFATc2 and NFATc4) is induced upon adipogenesis and in obese mice. Mice with the Nfatc2 ؊/؊ Nfatc4 ؊/؊ compound disruption exhibit defects in fat accumulation and are lean. Nfatc2 ؊/؊ Nfatc4 ؊/؊ mice are also protected from diet-induced obesity. Ablation of NFATc2 and NFATc4 increases insulin sensitivity, in part, by sustained activation of the insulin signaling pathway. Nfatc2 ؊/؊ Nfatc4 ؊/؊ mice also exhibit an altered adipokine profile, with reduced resistin and leptin levels. Mechanistically, NFAT is recruited to the transcription loci and regulates resistin gene expression upon insulin stimulation. Together, these results establish a role for NFAT in glucose/insulin homeostasis and expand the repertoire of NFAT function to metabolic pathogenesis and adipokine gene transcription.