Nitrite is the driver, phytohormones are modulators while NO and H2O2act as promoters of NO2-induced cell death

This study aimed to understand the molecular mechanisms of nitrogen dioxide (NO 2 )-induced toxicity and cell death in plants. Exposure of Arabidopsis to high concentrations of NO 2 induced cell death in a dose-dependent manner. No leaf symptoms were visible after fumigation for 1 h with 10 parts per million (ppm) NO 2 However, 20 ppm NO 2 caused necrotic lesion formation and 30 ppm NO 2 complete leaf collapse, which had already started during the 1 h fumigation period. NO 2 fumigation resulted in a massive accumulation of nitrite and in protein modifications by S-nitrosylation and tyrosine nitration. Nitric oxide (NO) at 30 ppm did not trigger leaf damage or any of the effects observed after NO 2 fumigation. The onset of NO 2 -induced cell death correlated with NO and hydrogen peroxide (H 2 O 2 ) signaling and a decrease in antioxidants. NO- and H 2 O 2 -accumulating mutants were more sensitive to NO 2 han wild-type plants. Accordingly, experiments with specific scavengers confirmed that NO and H 2 O 2 are essential promoters of NO 2 -induced cell death. Leaf injection of 100 mM nitrite caused an increase in S-nitrosylation, NO, H 2 O 2 , and cell death suggesting that nitrite functioned as a mediator of NO 2 -induced effects. A targeted screening of phytohormone mutants revealed a protective role of salicylic acid (SA) signaling in response to NO 2 It was also shown that phytohormones were modulators rather than inducers of NO 2 -induced cell death. The established experimental set-up is a suitable system to investigate NO 2 and cell death signaling in large-scale mutant screens.