Involvement of the leaf-specific multidrug and toxic compound extrusion (mate) transporter Nt-JAT2 in vacuolar sequestration of nicotine in nicotiana tabacum

Nobukazu Shitan, Shota Minami, Masahiko Morita, Minaho Hayashida, Shingo Ito, Kojiro Takanashi, Hiroshi Omote, Yoshinori Moriyama, Akifumi Sugiyama, Alain Goossens, Masataka Moriyasu, Kazufumi Yazaki

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Abstract

Alkaloids play a key role in higher plant defense against pathogens and herbivores. Following its biosynthesis in root tissues, nicotine, the major alkaloid of Nicotiana species, is translocated via xylem transport toward the accumulation sites, leaf vacuoles. Our transcriptome analysis of methyl jasmonate-treated tobacco BY-2 cells identified several multidrug and toxic compound extrusion (MATE) transporter genes. In this study, we characterized a MATE gene, nicotiana tabacum¿asmonateinducible alkaloid transporter 2 (Nt-JAT2), which encodes a protein that has 32% amino acid identity with Nt-JATI. Nt-JAT2 mRNA is expressed at a very low steady state level in whole plants, but is rapidly upregulated by methyl jasmonate treatment in a leaf-specific manner. To characterize the function of Nt-JAT2, yeast cells were used as the host organism in a cellular transport assay. Nt-JAT2 was localized at the plasma membrane In yeast cells. When incubated in nicotinecontaining medium, the nicotine content in Nt-JAT2-expressing cells was significantly lower than in control yeast. Nt-JAT2-expressing cells also showed lower content of other alkaloids like anabasine and anatabine, but not of flavonoids, suggesting that NI-JAT2 transports various alkaloids including nicotine. Fluorescence assays in BY-2 cells showed that Nt-JAT2-GFP was localized to the tonoplast. These findings indicate that Nt-JAT2 is involved in nicotine sequestration in leaf vacuoles following the translocation of nicotine from root tissues.

Original languageEnglish
Article numbere108789
JournalPloS one
Volume9
Issue number9
DOIs
Publication statusPublished - Sep 30 2014
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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    Shitan, N., Minami, S., Morita, M., Hayashida, M., Ito, S., Takanashi, K., Omote, H., Moriyama, Y., Sugiyama, A., Goossens, A., Moriyasu, M., & Yazaki, K. (2014). Involvement of the leaf-specific multidrug and toxic compound extrusion (mate) transporter Nt-JAT2 in vacuolar sequestration of nicotine in nicotiana tabacum. PloS one, 9(9), [e108789]. https://doi.org/10.1371/journal.pone.0108789