Phytodetoxification of TNT by transplastomic tobacco (Nicotiana tabacum) expressing a bacterial nitroreductase

Abstract

Expression of the bacterial nitroreductase gene, nfsI, in tobacco plastids conferred the ability to detoxify TNT. The toxic pollutant 2,4,6-trinitrotoluene (TNT) is recalcitrant to degradation in the environment. Phytoremediation is a potentially low cost remediation technique that could be applied to soil contaminated with TNT; however, progress is hindered by the phytotoxicity of this compound. Previous studies have demonstrated that plants transformed with the bacterial nitroreductase gene, nfsI have increased ability to tolerate and detoxify TNT. It has been proposed that plants engineered to express nfsI could be used to remediate TNT on military ranges, but this could require steps to mitigate transgene flow to wild populations. To address this, we have developed nfsI transplastomic tobacco (Nicotiana tabacum L.) to reduce pollen-borne transgene flow. Here we have shown that when grown on solid or liquid media, the transplastomic tobacco expressing nfsI were significantly more tolerant to TNT, produced increased biomass and removed more TNT from the media than untransformed plants. Additionally, transplastomic plants expressing nfsI regenerated with high efficiency when grown on medium containing TNT, suggesting that nfsI and TNT could together be used to provide a selectable screen for plastid transformation.

Evolutionary aspects of the Viridiplantae nitroreductases

“…Most probably, plant nitroreductases are participating in the oxidative stress, pollutant, and herbicide responses [7,65]. Also, some connections to circadian rhythms or the efficiency of the photosynthetic machine are possible.…”

“…Bacterial enzymes are not much suitable for such purposes, because they lack membrane anchor and their localization is not specific. Partially, this problem could be solved by transplastomic transformation, providing production of the high amount of functional enzyme [65]. From the other side, this method has several limitations, mainly: (1) many pollutants are absorbed by roots, where chloroplastic enzymes are not presented, or their activity is very low [11]; (2) plastid genes are greatly downregulated in fruits, where pollutants are often concentrated [29]; in general, plastid transformation is well-established only in the limited number of species, but agricultural and industrial plants species are rather recalcitrant [9].…”

Genetic modification of western wheatgrass (Pascopyrum smithii) for the phytoremediation of RDX and TNT

“…These transgenic plants were all able to degrade RDX and resist high levels of TNT when compared to 85 wild-type plants. In our recent report, nfsI transformed into the plastid genome of tobacco conferred enhanced resistance to TNT (Zhang et al 2017b). However, Arabidopsis, tobacco, and creeping bentgrass are not well-adapted to thriving in the challenging environments found in military training ranges.…”

Mechanism of Two-/Four-Electron Reduction of Nitroaromatics by Oxygen-Insensitive Nitroreductases: The Role of a Non-Enzymatic Reduction Step

“…The two-electron reduction of nitroaromatics by bacterial nitroreductases (NRs) or mammalian DT-diaphorase results in the formation of nitroso-(Ar-NO), and, subsequently, hydroxylamine (Ar-NHOH) products ( Scheme 1 ), which alkylate DNA and other biomolecules [ 2 ]. Apart from their role in the cytotoxicity of nitroaromatics, the latter reactions are important in the biodegradation of toxic environmental pollutants such as the polynitroaromatic explosives 2,4,6-trinitrotoluene (TNT) or 2,4,6-trinitrophenyl- N -methylnitramine (tetryl) [ 3 , 4 ].…”