Abstract
Melatonin was discovered in plants more than two decades ago and, especially in the last decade, it has captured the interests of plant biologists. Beyond its possible participation in photoperiod processes and its role as a direct free radical scavenger as well as an indirect antioxidant, melatonin is also involved in plant defense strategies/reactions. However, the mechanisms that this indoleamine activates to improve plant stress tolerance still require identification and clarification. In the present report, the ability of exogenous melatonin to protect Nicotiana tabacum L. line Bright Yellow 2 (BY-2) suspension cells against the toxic exposure to lead was examined. Studies related to cell proliferation and viability, DNA fragmentation, possible translocation of cytochrome c from mitochondria to cytosol, cell morphology after fluorescence staining and also the in situ accumulation of superoxide radicals measured via the nitro blue tetrazolium reducing test, were conducted. This work establishes a novel finding by correcting the inhibition of release of mitochondrial ctytocrome c in to the cytoplasm with the high accumulation of superoxide radicals. The results show that pretreatment with 200 nm of melatonin protected tobacco cells from DNA damage caused by lead. Melatonin, as an efficacious antioxidant, limited superoxide radical accumulation as well as cytochrome c release thereby, it likely prevents the activation of the cascade of processes leading to cell death. Fluorescence staining with acridine orange and ethidium bromide documented that lead-stressed cells additionally treated with melatonin displayed intact nuclei. The results revealed that melatonin at proper dosage could significantly increase BY-2 cell proliferation and protected them against death. It was proved that melatonin could function as an effective priming agent to promote survival of tobacco cells under harmful lead-induced stress conditions.
Highlights
In their natural environment, plants are exposed to many different biotic and abiotic stresses
The experiments were performed in four variants: (i) C: Bright Yellow 2 (BY-2) cells cultured under optimal conditions on LS medium, (ii) MEL: BY-2 cells cultured on LS medium supplemented with melatonin from the start of new culture; (iii) Pb: BY-2 cells cultured on LS medium with Pb2+ added on the 4th day of culture and (iv) MEL + Pb: BY-2 cells cultured on LS medium with melatonin added from the start of culture and stressed with Pb2+ added on the 4th day of culture
From the first day after heavy metal stress induction, proliferation of the MEL + Pb cells was about 40% higher in comparison to those treated with Pb but not primed with melatonin (Pb) (Figure 1A) this tendency was maintained throughout the duration of the Pb-stress
Summary
Plants are exposed to many different biotic and abiotic stresses. Heavy metals, especially lead (Pb), are major environmental pollutants, in areas with high anthropogenic pressure (Gill, 2014) and its accumulation has adverse effects on plant growth and crop productivity. The level of heavy metals in agricultural soil is normally very low, the repeated use of phosphate fertilizers over long periods may cause dangerously high concentrations of some of these toxins (Gill, 2014). Plant roots are not selective and absorb Pb with other minerals where accumulates. In a number of species, high Pb levels cause abnormal plant morphology, reduced plant growth and it induces cell death (Pourrut et al, 2012). Pb provokes DNA damage, gene mutations, protein oxidation, lipid peroxidation and it promotes signal transduction cascades that promote cell death (Wierzbicka, 1999; Gill, 2014)
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