Dual NADPH oxidases DUOX1 and DUOX2 synthesize NAADP and are necessary for Ca2+ signaling during T cell activation

An origin story for NAADP in T cellsEarly steps in T cell activation are mediated by the synthesis of Ca2+-mobilizing second messenger NAADP, which is produced through oxidation of NAADPH by a previously unknown enzyme. Gu et al. identified NAADPH-oxidizing enzymes that were critical for the early phases of T cell activation. Stimulation of T cell receptors initially results in rapid production of NAADP that induces the formation of localized Ca2+ microdomains that eventually lead to more global and sustained intracellular Ca2+ signaling. In cultured rat T cells, knockout of DUOX2 reduced local Ca2+ microdomain formation, whereas functional knockout of both DUOX1 and DUOX2 in murine T cells suppressed global intracellular Ca2+ signaling. The findings identify a critical pair of enzymes in T cell activation.The formation of Ca2+ microdomains during T cell activation is initiated by the production of nicotinic acid adenine dinucleotide phosphate (NAADP) from its reduced form NAADPH. The reverse reaction—NAADP to NAADPH—is catalyzed by glucose 6-phosphate dehydrogenase (G6PD). Here, we identified NADPH oxidases NOX and DUOX as NAADP-forming enzymes that convert NAADPH to NAADP under physiological conditions in vitro. T cells express NOX1, NOX2, and, to a minor extent, DUOX1 and DUOX2. Local and global Ca2+ signaling were decreased in mouse T cells with double knockout of Duoxa1 and Duoxa2 but not with knockout of Nox1 or Nox2. Ca2+ microdomains in the first 15 s upon T cell activation were significantly decreased in Duox2−/− but not in Duox1−/− T cells, whereas both DUOX1 and DUOX2 were required for global Ca2+ signaling between 4 and 12 min after stimulation. Our findings suggest that a DUOX2- and G6PD-catalyzed redox cycle rapidly produces and degrades NAADP through NAADPH as an inactive intermediate.