Supplementary Figure 1: TNT reduces cellular NAD⁺ levels by degradation.

(a) Global gene expression after induction of tnt in E. coli was determined by deep sequencing. Gene expression changes were compared to E. coli containing an empty vector and are plotted as a ratio (log₂-fold changes) against the significance of the difference in expression (Log Odds; −log₁₀ q-value). Only genes with expression ratios larger than four-fold (log₂ ratios ≥ 2 or ≤ −2) and log Odds > 1.3 (q-values < 0.05) were included. (b) The amount of NAD⁺ extracted from 5 × 10⁸ E. coli or 1 × 10⁶ Jurkat cells 2 h or 24 h post-induction of tnt expression, respectively, was measured by a colorimetric enzyme coupling assay. Data represent the means ± s.d. of three independent experiments. (c) NAD⁺ hydrolysis by TNT releases ADP-ribose (and nicotinamide) as determined by thin-layer chromatography using radiolabeled NAD⁺, cyclic ADP-ribose, and ADP-ribose as standards as described in Methods.