NFAT control of immune function: New Frontiers for an Abiding Trooper

Abstract

Nuclear factor of activated T cells (NFAT) was first described almost three decades ago as a Ca ²⁺/calcineurin-regulated transcription factor in T cells. Since then, a large body of research uncovered the regulation and physiological function of different NFAT homologues in the immune system and many other tissues. In this review, we will discuss novel roles of NFAT in T cells, focusing mainly on its function in humoral immune responses, immunological tolerance, and the regulation of immune metabolism.

Keywords: CRAC; Calcium; NFAT; STIM; T cell; anergy; exhaustion; metabolism.

Figure 1.. Canonical and alternative NFAT activation in T cells

( A) Antigen receptor stimulation leads to the production of inositol-1,4,5-trisphosphate (IP ₃), which opens IP ₃ receptor channels in the endoplasmic reticulum (ER). The subsequent decrease in the ER Ca ²⁺ concentration activates stromal interaction molecule 1 (STIM1) and STIM2, which then bind to and open Ca ²⁺ release-activated Ca ²⁺ (CRAC) channels formed by ORAI1 and ORAI2 proteins in the plasma membrane. Ca ²⁺ influx through CRAC channels activates calmodulin (CaM) and the serine/threonine phosphatase calcineurin. Calcineurin dephosphorylates multiple serine/threonine residues in the regulatory domain of NFAT, resulting in a conformational change, exposure of nuclear localization signals, and nuclear import of NFAT. ( B) Janus kinase 3 (Jak3) downstream of the interleukin-7 (IL-7) receptor phosphorylates a single tyrosine residue within the regulatory domain of NFAT2, which induces nuclear translocation and activation of NFAT2 independent of Ca ²⁺ signals and calcineurin in thymocytes. CsA, cyclosporine A; NFAT, nuclear factor of activated T cells.

Figure 2.. Distinct NFAT functions in effector, follicular, regulatory, and tolerized T cells.

( A) Stimulation of conventional T cells via the T-cell receptor (TCR) and co-stimulatory receptors results in NFAT activation and its cooperative DNA binding together with AP-1 (Jun/Fos). NFAT:AP-1 complexes regulate the expression of cytokines and other effector molecules as well as the short NFAT2/αA isoform that further enhances T-cell activation in a positive auto-regulatory loop. ( B) In T follicular helper (Tfh) cells, NFAT, together with AP-1, controls the expression of many genes that regulate the differentiation of Tfh cells (IRF4 and BATF), Tfh cell homing to B-cell follicles (CXCR5), and Tfh cell help to B cells (CD40L, IL-4, and IL-21). ( C) In peripherally induced regulatory T (iTreg) cells, TCR-dependent NFAT and TGFβ-dependent Smad3 activation converge at the conserved non-coding sequence (CNS) 1 of the Foxp3 locus to induce Foxp3 expression. ( D) In thymus-derived “natural” Treg (nTreg) cells, Foxp3 expression appears largely independent of NFAT activation. In both iTreg and nTreg cells, NFAT forms a ternary NFAT:Foxp3 complex with DNA that induces the expression of Treg-associated genes such as CD25 and CTLA-4 and antagonizes the expression of pro-inflammatory genes and the short NFAT2/αA isoform. In follicular Treg (Tfr) cells, NFAT regulates the expression of CXCR5, CTLA-4, and PD-1 that are required for Tfr cell function. ( E, F) Chronic TCR stimulation without co-stimulation triggers the formation of NFAT homomeric complexes that induce a gene expression program associated with T-cell anergy ( E) or exhaustion ( F) and that is distinct from NFAT:AP-1 complex-mediated gene expression. ( E) NFAT-dependent genes associated with anergic CD4 ⁺ T cells include E3 ubiquitin ligases (GRAIL, Itch, and Cbl-b) and caspase 3 that target molecules involved in proximal TCR signaling, which renders T cells unresponsive to re-stimulation. ( F) NFAT-dependent genes associated with exhausted CD8 ⁺ T cells are similar to those in anergic CD4 ⁺ T cells but also include inhibitory receptors such as PD-1, TIM3, and LAG3 that antagonize TCR signaling. NFAT, nuclear factor of activated T cells; TGFβ, transforming growth factor beta.

Figure 3.. NFAT regulates T-cell metabolism.

Ca ²⁺/calcineurin/NFAT signaling following T-cell receptor (TCR) and co-stimulation is required for the switch from catabolic to high-rate anabolic metabolism of activated T cells, cell cycle entry, and proliferation. NFAT directly controls the expression of “metabolic master regulators” such as IRF4, HIF1α, and potentially c-Myc that subsequently induce the expression of glucose transporter 1 (GLUT1), glycolytic enzymes, and mitochondrial electron transport chain (ETC) complexes that mediate aerobic glycolysis and mitochondrial respiration, respectively. In addition, NFAT directly controls the expression of the high-affinity glucose transporter GLUT3. NFAT also senses the metabolic state of T cells. The glycolytic intermediate phosphoenolpyruvate (PEP) inhibits the sarco/endoplasmic reticulum Ca ²⁺ ATPase (SERCA) and thereby enhances Ca ²⁺ signaling and NFAT activation, whereas lactate inhibits NFAT activation. Low levels of reactive oxygen species (ROS) generated by mitochondrial respiration enhance NFAT activation, whereas high ROS levels inhibit NFAT. CRAC, Ca ²⁺ release-activated Ca ²⁺; CsA, cyclosporine A; ER, endoplasmic reticulum; IP ₃, inositol-1,4,5-tisphosphate; NFAT, nuclear factor of activated T cells; STIM, stromal interaction molecule.