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Plant Molecular Biology

, Volume 88, Issue 1–2, pp 119–131 | Cite as

The cytosolic branched-chain aminotransferases of Arabidopsis thaliana influence methionine supply, salvage and glucosinolate metabolism

  • Kurt Lächler
  • Janet Imhof
  • Michael Reichelt
  • Jonathan Gershenzon
  • Stefan Binder
Article

Abstract

Arabidopsis thaliana possesses six branched-chain aminotransferases (BCAT1–6). Previous studies revealed that some members of this protein family are involved in the biosynthesis of branched-chain amino acids and/or in the Met chain elongation pathway, the initial steps towards the biosynthesis of Met-derived glucosinolates. We now analyzed branched-chain aminotransferase 6 (BCAT6). In vivo GFP-tagging experiments strongly suggest this enzyme to be localized to the cytosol. Substrate specificity assays performed with recombinant enzyme revealed that BCAT6 transaminates Val, Leu and Ile as well as the corresponding 2-oxo acids but also transaminates Met and its cognate ketoacid 4-methyl-2-oxobutanoate. We established single (bcat6-1), double (bcat4-2/bcat6-1) and triple (bcat3-1/bcat4-2/bcat6-1) mutants involving BCAT6 with the latter exhibiting a clear macroscopic phenotype with smaller plants and abnormal leaf morphology. Metabolite profiling of these mutants demonstrated that BCAT6 can contribute to Met chain elongation with the triple mutant line lacking BCAT3, 4 and 6 showing a dramatic reduction of Met-derived glucosinolate species down to 32 and 14 % of wild-type levels in plant foliage and seeds, respectively. This drop in glucosinolate levels is accompanied by a 46-fold increase of free Met, demonstrating the important role of the three branched-chain aminotransferases in converting Met to its 2-oxo acid for glucosinolate chain elongation. In addition, we determined the relative amounts of 5′-deoxy-5′-methylthioadenosine, an intermediate of the Met recycling pathway. This metabolite accumulated to relative high amounts in the absence of the cytosolic BCAT4 and BCAT6, suggesting that cytosolic Met salvage also contributes to the biosynthesis of glucosinolates.

Keywords

Arabidopsis thaliana Amino acids Glucosinolates Branched-chain aminotransferases 

Notes

Acknowledgments

We thank Ulrike Tengler and Conny Guha for excellent technical assistance. We are also grateful to Laura Maccarone and Sarah Kovarik for their engagement in subcellular localization studies. This work was supported by Grant Bi 590/9-2 from the Deutsche Forschungsgemeinschaft.

Supplementary material

11103_2015_312_MOESM1_ESM.pdf (166 kb)
Supplementary material 1 (PDF 167 kb)
11103_2015_312_MOESM2_ESM.pdf (935 kb)
Supplementary material 2 (PDF 936 kb)
11103_2015_312_MOESM3_ESM.pdf (92 kb)
Supplementary material 3 (PDF 93 kb)

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Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Kurt Lächler
    • 1
  • Janet Imhof
    • 1
  • Michael Reichelt
    • 2
  • Jonathan Gershenzon
    • 2
  • Stefan Binder
    • 1
  1. 1.Institut Molekulare BotanikUniversität UlmUlmGermany
  2. 2.Abteilung BiochemieMax Planck Institut für Chemische ÖkologieJenaGermany

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