Expression of a constitutively active nitrate reductase variant in tobacco reduces tobacco‐specific nitrosamine accumulation in cured leaves and cigarette smoke

• Published in: Plant biotechnology journal Vol. 14; no. 7; pp. 1500 - 1510
• Main Authors: Lu, Jianli, Zhang, Leichen, Lewis, Ramsey S., Bovet, Lucien, Goepfert, Simon, Jack, Anne M., Crutchfield, James D., Ji, Huihua, Dewey, Ralph E.
• Format: Journal Article
• Language: English
• Published: England John Wiley & Sons, Inc 01.07.2016; John Wiley and Sons Inc
• Subjects: Accumulation; Amino acids; Ammonia; burley; Carcinogens; Carcinogens - metabolism; Chlorophyll; Cigarette smoke; Cigarettes; Dehydrogenases; Enzymes; Genes; Genetic engineering; Genetic research; Genotype & phenotype; Laboratories; Leaves; Metabolic Networks and Pathways; Metabolites; Mutation; N'-Nitrosonornicotine; Nicotiana - genetics; Nicotiana - metabolism; Nicotiana tabacum; Nicotine; Nitrate reductase; Nitrate Reductase - genetics; Nitrates; Nitrates - metabolism; Nitrogen; Nitrogen - metabolism; nitrogen‐assimilation pathway; Nitrosamine; Nitrosamines; Nitrosamines - metabolism; NNK; NNN; Phenotypes; Plant Leaves - genetics; Plant Leaves - metabolism; Plants; Plants, Genetically Modified - metabolism; Protocol; Reductases; Smoke; Smoking; Tobacco; Tobacco Products; tobacco‐specific nitrosamines; North Carolina; United States > US; burley; tobacco-specific nitrosamines; NNK; nitrate reductase; tobacco; nitrogen-assimilation pathway; NNN
• ISSN: 1467-7644; 1467-7652
• DOI: 10.1111/pbi.12510

Summary Burley tobaccos (Nicotiana tabacum) display a nitrogen‐use‐deficiency phenotype that is associated with the accumulation of high levels of nitrate within the leaf, a trait correlated with production of a class of compounds referred to as tobacco‐specific nitrosamines (TSNAs). Two TSNA species, 4‐(methylnitrosamino)‐1‐(3‐pyridyl)‐1‐butanone (NNK) and N‐nitrosonornicotine (NNN), have been shown to be strong carcinogens in numerous animal studies. We investigated the potential of molecular genetic strategies to lower nitrate levels in burley tobaccos by overexpressing genes encoding key enzymes of the nitrogen‐assimilation pathway. Of the various constructs tested, only the expression of a constitutively active nitrate reductase (NR) dramatically decreased free nitrate levels in the leaves. Field‐grown tobacco plants expressing this NR variant exhibited greatly reduced levels of TSNAs in both cured leaves and mainstream smoke of cigarettes made from these materials. Decreasing leaf nitrate levels via expression of a constitutively active NR enzyme represents an exceptionally promising means for reducing the production of NNN and NNK, two of the most well‐documented animal carcinogens found in tobacco products.