Reactive nitrogen species (RNS) play an important role in plant immunity as signaling factors. We previously developed a plasma technology to partially convert air molecules into dinitrogen pentoxide (N2O5), an RNS whose physiological action is poorly understood. To reveal the function of N2O5 gas in plant immunity, Arabidopsis thaliana was exposed to plasma-generated N2O5 gas once (20 s) per day for 3 days, and inoculated with Botrytis cinerea, Pseudomonas syringae pv. tomato DC3000 (Pst), or cucumber mosaic virus strain yellow (CMV(Y)) at 24 h after the final N2O5 gas exposure. Lesion size with B. cinerea infection was significantly (P < 0.05) reduced by exposure to N2O5 gas. Propagation of CMV(Y) was suppressed in plants exposed to N2O5 gas compared with plants exposed to the air control. However, proliferation of Pst in the N2O5-gas-exposed plants was almost the same as in the air control plants. These results suggested that N2O5 gas exposure could control plant disease depending on the type of pathogen. Furthermore, changes in gene expression at 24 h after the final N2O5 gas exposure were analyzed by RNA-Seq. Based on the gene ontology analysis, jasmonic acid and ethylene signaling pathways were activated by exposure of Arabidopsis plants to N2O5 gas. A time course experiment with qRT-PCR revealed that the mRNA expression of the transcription factor genes, WRKY25, WRKY26, WRKY33, and genes for tryptophan metabolic enzymes, CYP71A12, CYP71A13, PEN2, and PAD3, was transiently induced by exposure to N2O5 gas once for 20 s peaking at 1–3 h post-exposure. However, the expression of PDF1.2 was enhanced beginning from 6 h after exposure and its high expression was maintained until 24–48 h later. Thus, enhanced tryptophan metabolism leading to the synthesis of antimicrobial substances such as camalexin and antimicrobial peptides might have contributed to the N2O5-gas-induced disease resistance.