Chemotaxis of Mouse Bone Marrow Neutrophils and Dendritic Cells Is Controlled by ADP-Ribose, the Major Product Generated by the CD38 Enzyme Reaction

Abstract

The ectoenzyme CD38 catalyzes the production of cyclic ADP-ribose (cADPR) and ADP-ribose (ADPR) from its substrate, NAD+. Both products of the CD38 enzyme reaction play important roles in signal transduction, as cADPR regulates calcium release from intracellular stores and ADPR controls cation entry through the plasma membrane channel TRPM2. We previously demonstrated that CD38 and the cADPR generated by CD38 regulate calcium signaling in leukocytes stimulated with some, but not all, chemokines and controls leukocyte migration to inflammatory sites. However, it is not known whether the other CD38 product, ADPR, also regulates leukocyte trafficking In this study we characterize 8-bromo (8Br)-ADPR, a novel compound that specifically inhibits ADPR-activated cation influx without affecting other key calcium release and entry pathways. Using 8Br-ADPR, we demonstrate that ADPR controls calcium influx and chemotaxis in mouse neutrophils and dendritic cells activated through chemokine receptors that rely on CD38 and cADPR for activity, including mouse FPR1, CXCR4, and CCR7. Furthermore, we show that the calcium and chemotactic responses of leukocytes are not dependent on poly-ADP-ribose polymerase 1 (PARP-1), another potential source of ADPR in some leukocytes. Finally, we demonstrate that NAD+ analogues specifically block calcium influx and migration of chemokine-stimulated neutrophils without affecting PARP-1-dependent calcium responses. Collectively, these data identify ADPR as a new and important second messenger of mouse neutrophil and dendritic cell migration, suggest that CD38, rather than PARP-1, may be an important source of ADPR in these cells, and indicate that inhibitors of ADPR-gated calcium entry, such as 8Br-ADPR, have the potential to be used as anti-inflammatory agents.

NAD+ Levels Control Ca2+ Store Replenishment and Mitogen-induced Increase of Cytosolic Ca2+ by Cyclic ADP-ribose-dependent TRPM2 Channel Gating in Human T Lymphocytes

“…Production of intracellular ADPR is due to the enhanced NAD ϩ -glycohydrolase activity of CD38 in the presence of an increased concentration of its substrate NAD ϩ . Although CD38 is an ectoenzyme, with the active site facing the extracellular environment (59,60), its expression/activation in cells leads to the intracellular accumulation of cADPR, the product of its ADP-ribosyl cyclase activity (20,(61)(62)(63), and of ADPR, the product of its NAD ϩ -glycohydrolase activity (16,45).…”

“…In addition, TRPM2-regulated functions have been reported in different types of immune cells. In T lymphocytes, ADPR-mediated gating of TRPM2 has been implicated in the regulation of the [Ca 2ϩ ] i increase and subsequent cell death induced by high concentrations of ConA (16); in B lymphocytes, ADPR-mediated gating of TRPM2 occurs in response to oxidative stress (64); in neutrophils, the bacterial peptide formyl-methionyl-leucyl-phenylalanine (fMLP)-induced Ca 2ϩ influx is regulated by a signaling pathway involving CD38-ADPR-TRPM2 (45,65). Thus, increasing evidence points to TRPM2 as a central player for proper immune cell activity.…”

“…ADPR is a known TRPM2 agonist (28,(43)(44)(45). To assess a potential role for TRPM2 in the NAM-or NA-induced increase of Ca 2ϩ content in the TG-sensitive Ca 2ϩ stores, the expression of this channel was down-regulated by transfecting a specific siRNA.…”

Synergistic regulation of endogenous TRPM2 channels by adenine dinucleotides in primary human neutrophils

“…Thus, signaling pathways that cause peroxide, cADPR or NAADP generation must be considered regulators of TRPM2. In addition, these agonists synergize with reactive oxygen species, which play an important role in neutrophil function [41] and leukocyte chemotaxis [33,42].…”

Altered functional properties of a TRPM2 variant in Guamanian ALS and PD

“…TRPM2 is highly expressed in the brain, in both microglia and neuronal cells, but its biological role in these cells still needs to be understood. Studies suggest that Ca 2ϩ influx via TRPM2 is necessary for microglia and other phagocytes to mount effective inflammatory and clearance responses (10,29). An important advance in our understanding of the biological role of TRPM2 is provided by a recent study on TRPM2-KO mice (30).…”