Journal Article

Non-Functionalization of Two <i>CYP82E</i> Nicotine <i>N</i>-Demethylase Genes Abolishes Nornicotine Formation in <i>Nicotiana langsdorffii</i>

Phattharaporn Pakdeechanuan, Seddon Teoh, Tsubasa Shoji and Takashi Hashimoto

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 53, issue 12, pages 2038-2046
Published in print December 2012 | ISSN: 0032-0781
Published online October 2012 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcs139
Non-Functionalization of Two CYP82E Nicotine N-Demethylase Genes Abolishes Nornicotine Formation in Nicotiana langsdorffii

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Nornicotine is formed from nicotine by nicotine N-demethylase, a CYP82E family monooxygenase, and accumulates to high levels in some tobacco (Nicotiana tabacum) cultivars and many wild Nicotiana species. Nicotiana langsdorffii does not form nornicotine, whereas the closely related species N. alata accumulates this alkaloid abundantly. We show here that the two nicotine N-demethylase genes in N. langsdorffii have been inactivated by different molecular mechanisms. We identified four N. alata CYP82E genes that encode functional nicotine N-demethylases. In N. langsdorffii, however, one CYP82E gene encoding a functional enzyme was not expressed at all, whereas the other was weakly expressed but contained a one-nucleotide deletion in the first exon, yielding a truncated protein. Expression analysis of interspecific F1 hybrids between N. alata and N. langsdorffii indicated that cis-acting polymorphisms abolish expression of the otherwise functional CYP82E gene in N. langsdorffii. Segregation analysis of tobacco alkaloids and individual CYP82E alleles in F2 progeny revealed that duplicated CYP82E genes in both species are genetically linked, and provide genetic evidence that CYP82E genes are solely responsible for nornicotine formation in these wild Nicotiana species.

Keywords: CYP82; Nicotiana langsdorffii; Nicotine N-demethylase; Nornicotine

Journal Article.  5412 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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