Running Head: 2ʹ-Deoxy-ADPR is an endogenous superagonist of TRPM2

17 Transient receptor potential melastatin 2 (TRPM2), is a ligand-gated Ca-permeable non18 selective cation channel. While physiological stimuli, e.g. chemotactic agents, evoke 19 controlled Ca signals via TRPM2, pathophysiological signals, such as reactive oxygen 20 species or genotoxic stress result in prolonged TRPM2-mediated Ca entry and consequently 21 apoptosis. To date, adenosine 5ʹ-diphosphoribose (ADPR, 1) has been assumed to be the main 22 agonist for TRPM2. Here, we show that 2ʹ-deoxy-ADPR 2 was a significantly better TRPM2 23 2 agonist, inducing 10.4-fold higher whole cell currents at saturation. Mechanistically, this 24 increased activity was caused by decreased rate of inactivation and higher average open 25 probability. Using high performance liquid chromatography (HPLC) and mass spectrometry, 26 endogenous 2ʹ-deoxy-ADPR was detected in Jurkat T-lymphocytes. Consistently, cytosolic 27 nicotinamide mononucleotide adenylyltransferase 2 (NMNAT-2) and nicotinamide adenine 28 dinucleotide (NAD)-glycohydrolase CD38 sequentially catalyzed synthesis of 2ʹ-deoxy29 ADPR from nicotinamide mononucleotide and 2ʹ-deoxy-ATP in vitro. Thus, 2ʹ-deoxy-ADPR 30 is an endogenous TRPM2 superagonist that may act as cell signaling molecule. 31