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

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.

Genome‐wide association analysis reveals distinct genetic architectures for single and combined stress responses in Arabidopsis thaliana

“…Another QTL, identified for the response to P. rapae, contained, as most likely candidates, BCAT-1 and BCAT-2, which are enzymes involved in branched amino acid (leucine (Leu), valine (Val) and isoleucine (Ile)) (BCAA) metabolism (Diebold et al, 2002). Interestingly, homologues of these genes (BCAT-3, BCAT-4, BCAT-6) have been implicated in the production of aliphatic glucosinolates (Schuster et al, 2006;Lachler et al, 2015). Furthermore, co-expression networks have revealed that BCAT-4 is co-expressed with MYB29, MYB28 and several putative genes involved in Leu metabolism (Hirai et al, 2007).…”

Carbohydrate and amino acids metabolic response to heat stress in the intestine of the sea cucumberApostichopus japonicus

“…Valine, leucine and isoleucine are called branched-chain amino acids (BCAAs), characterized by their small branched hydrocarbon residues. The breakdown or synthesis of BCAAs depends on the catalysis of the family of branched-chain aminotransferases (BCATs) (Lachler, Imhof, Reichelt, Gershenzon & Binder, 2015). BCATs catalyse the crucial step in BCAAs metabolism, a reversible transamination from BCAAs and a-ketoglutarate to corresponding branched-acid a-keto acids and glutamate (Brosnan & Brosnan, 2006).…”

Decision letter: Independent evolution of ancestral and novel defenses in a genus of toxic plants (Erysimum, Brassicaceae)

“…Elbtalaue, but also identified additional aliphatic and indole glucosinolates at lower concentrations. By making use of the known glucosinolate biosynthetic genes from Arabidopsis (Hull et al 2000, Mikkelsen et al 2000, Kliebenstein et al 2001b, Kroymann et al 2001, Reintanz et al 2001, Chen et al 2003, Kroymann et al 2003, Grubb et al 2004, Mikkelsen et al 2004, Piotrowski et al 2004, Textor et al 2004, Nozawa et al 2005, Klein et al 2006, Schuster et al 2006, Hansen et al 2007, Textor et al 2007, Knill et al 2008, Li et al 2008, Farquharson 2009, Geu-Flores et al 2009, Gigolashvili et al 2009, He et al 2009, Klein and Papenbrock 2009, Knill et al 2009, Pfalz et al 2009, Sawada et al 2009, He et al 2010, Geu-Flores et al 2011, He et al 2011, Pfalz et al 2011, Lachler et al 2015, Kong et al 2016, Pfalz et al 2016,…”