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Abstract
Lead is a heavy metal known to be toxic to both animals and plants. Nitric oxide (NO) was reported to participate in plant responses to different heavy metal stresses. In this study, we analyzed the function of exogenous and endogenous NO in Pb-induced toxicity in tobacco BY-2 cells, focusing on the role of NO in the generation of reactive oxygen species (ROS) as well as Pb2+ and Ca2+ fluxes using non-invasive micro-test technology (NMT). Pb treatment induced BY-2 cell death and rapid NO and ROS generation, while NO burst occurred earlier than ROS accumulation. The elimination of NO by 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) resulted in a decrease of ROS, and the supplementation of NO by sodium nitroprusside (SNP) caused an increased accumulation of ROS. Furthermore, the addition of exogenous NO stimulated Pb2+ influx, thus promoting Pb uptake in cells and aggravating Pb-induced toxicity in cells, whereas the removal of endogenous NO produced the opposite effect. Moreover, we also found that both exogenous and endogenous NO enhanced Pb-induced Ca2+ effluxes and calcium homeostasis disorder. These results suggest that exogenous and endogenous NO played a critical regulatory role in BY-2 cell death induced by Pb stress by promoting Pb2+ influx and accumulation and disturbing calcium homeostasis.
Highlights
Heavy metal contamination is a major form of environmental pollution owing to emissions from industry, agricultural chemicals, vehicular traffic, and other human activities [1]
The results presented here show that exogenous nitric oxide (NO) and Pb-triggered endogenous NO burst contributed to reactive oxygen species (ROS) generation in tobacco BY-2 cells, promoted Pb2+ influx in cells, and increased Pb uptake by the cells, enhanced Pb-induced calcium homeostasis disorder, and played a critical regulatory role in tobacco BY-2 cell death induced by Pb stress
The dead cells were determined by 3 μg/mL propidium iodide (PI), which is generally excluded from viable cells for its membrane-impermeable properties
Summary
Heavy metal contamination is a major form of environmental pollution owing to emissions from industry, agricultural chemicals, vehicular traffic, and other human activities [1]. Lead is one of the most hazardous metals in the environment and is toxic to plants and other organisms. Pb impairs plants by inhibiting seed germination and plant growth [2,3,4]; reducing nutrient uptake and biomass [5]; disrupting cell membrane permeability, photosynthesis, and cell division [4,6,7]; inhibiting fundamental enzymatic reactions; demolishing cell viability; and inducing cell death [8]. Heavy metals usually result in oxidative stress and a burst of reactive oxygen species (ROS) [9]. In addition to ROS, nitric oxide (NO) may be rapidly induced in plant cells to regulate plant responses to abiotic stress, including heavy metal toxicity [10,11].
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