Fig. 1

Induction of systemic resistance by exogenous NAD(P)⁺ and movement of exogenously applied NAD⁺. a, b NAD a and NADP b leakage from the wild-type Col-0 leaves infiltrated with 10 mm MgCl₂ (mock) or Psm (OD₆₀₀ = 0.002). One leaf disk was removed from each infiltrated leaf and sets of 10 leaf disks were submerged in 5 mL water in test tubes. NAD(P) concentrations in the water were measured over time by enzymatic cycling assays. Data represent the mean ± standard deviation (SD) of three biological replicates. Asterisks denote significant differences between Psm- and MgCl₂-treated samples (*p < 0.05, **p < 0.01; Student’s t test). c, d Expression of PR1 c and growth of Psm d in the systemic leaves of the wild-type Col-0 plants treated with MgCl₂, Psm, H₂O, NAD⁺, or NADP⁺. For MgCl₂ and Psm treatments, three lower leaves on each 4-week-old soil-grown Arabidopsis plant were infiltrated with 10 mm MgCl₂ or a Psm suspension (OD₆₀₀ = 0.002). Two days later, two systemic leaves were either collected for PR1 expression analysis by qPCR c or challenge-inoculated with Psm (OD₆₀₀ = 0.001) d. Three days later, eight leaves were collected to examine the growth of the pathogen. Alternatively, three lower leaves were infiltrated with H₂O, 0.4 mm NAD⁺, or 0.8 mm NADP⁺ every 12 hr for a total of four times. About 5 hr after the last infiltration, two systemic leaves were either collected for PR1 analysis c or challenge-inoculated with Psm (OD₆₀₀ = 0.001) d. Expression levels of PR1 c were normalized against the constitutively expressed UBQ5. Data represent the mean ± SD of three c or eight biological replicates d. Different letters denote significant differences (p < 0.05; one-way ANOVA with Tukey’s test). Compared with the strong systemic resistance induced by Psm (~ 35-fold decrease in Psm growth), NAD⁺ and NADP⁺ induced intermediate levels of resistance in the systemic leaves (~ 6.5-fold decrease in Psm growth). e, f Autoradiographic detection of ³²P in the systemic leaves of Arabidopsis e and N. benthamiana f plants with the lower leaves infiltrated with ³²P-NAD⁺. Three lower leaves on an Arabidopsis plant or two lower leaves on a N. benthamiana plant were infiltrated with a water solution of 6.25 nm ³²P-NAD⁺ plus 1 mm unlabeled NAD⁺. Twenty-four hr later, the infiltrated leaves (I in red) and two systemic leaves (U in blue) were collected and exposed to X-ray film