Characterization of the Branched-Chain Amino Acid Aminotransferase Enzyme Family in Tomato    

Abstract

Branched-chain amino acids (BCAAs) are synthesized in plants from branched-chain keto acids, but their metabolism is not completely understood. The interface of BCAA metabolism lies with branched-chain aminotransferases (BCAT) that catalyze both the last anabolic step and the first catabolic step. In this study, six BCAT genes from the cultivated tomato (Solanum lycopersicum) were identified and characterized. SlBCAT1, -2, -3, and -4 are expressed in multiple plant tissues, while SlBCAT5 and -6 were undetectable. SlBCAT1 and -2 are located in the mitochondria, SlBCAT3 and -4 are located in chloroplasts, while SlBCAT5 and -6 are located in the cytosol and vacuole, respectively. SlBCAT1, -2, -3, and -4 were able to restore growth of Escherichia coli BCAA auxotrophic cells, but SlBCAT1 and -2 were less effective than SlBCAT3 and -4 in growth restoration. All enzymes were active in the forward (BCAA synthesis) and reverse (branched-chain keto acid synthesis) reactions. SlBCAT3 and -4 exhibited a preference for the forward reaction, while SlBCAT1 and -2 were more active in the reverse reaction. While overexpression of SlBCAT1 or -3 in tomato fruit did not significantly alter amino acid levels, an expression quantitative trait locus on chromosome 3, associated with substantially higher expression of Solanum pennellii BCAT4, did significantly increase BCAA levels. Conversely, antisense-mediated reduction of SlBCAT1 resulted in higher levels of BCAAs. Together, these results support a model in which the mitochondrial SlBCAT1 and -2 function in BCAA catabolism while the chloroplastic SlBCAT3 and -4 function in BCAA synthesis.

Chilling-induced tomato flavor loss is associated with altered volatile synthesis and transient changes in DNA methylation

“…Isovaleraldehyde, 3-methyl-1-butanol, and 2-methyl-1-butanol are believed to be derived from the immediate precursors of branched-chain amino acids. Branched-chain aminotransferases (BCATs) catalyze the last step in synthesis and the first step in catabolism of these amino acids (23,24). Transcripts of two members of the BCAT gene family were significantly lower after 8 d of chilling and remained low after 1 d of recovery, in accord with the volatile contents (Fig.…”

Transcriptome and Metabolite Profiling Show That APETALA2a Is a Major Regulator of Tomato Fruit Ripening

“…The majority of the amino acids (15 out of 22) were present at significantly lower levels in the AP2i fruits than in the wild type, and the most dramatic reductions were in b-Ala, Ile, Met, Phe, and Trp. The observed reduced levels of Ile and Leu (branchedchain amino acids) are in agreement with the higher expression of BRANCHED-CHAIN AMINOTRANSFERASE1 (BCAT1; 8.6-fold), which was shown to be involved in the degradation of these two amino acids (Maloney et al, 2010). Similarly, nine of the 15 organic acids measured displayed significant changes.…”

Identification and Mode of Inheritance of Quantitative Trait Loci for Secondary Metabolite Abundance in Tomato

“…As mentioned above, the cloning of QTLs from this population remains an arduous task, and we have genetically confirmed only a handful of the candidate genes we identified on the basis of our GC-MS screening most notably those for branched-chain amino acids (Maloney et al, 2010) and tocopherol (Quadrana et al, 2013(Quadrana et al, , 2014. As a first step, given that secondary metabolism is predominantly under transcriptional control Patra et al, 2013;Tohge et al, 2014), we used a previously established quantitative realtime PCR platform to define differences in transcription factor expression in eight of the ILs, as well as defining the expression level of a range of genes encoding key enzymes of secondary metabolism.…”

“…In tomato, the majority of both natural variance and metabolite quantitative trait loci (mQTL) studies have focused on primary metabolism (Schauer et al, 2005(Schauer et al, , 2008Stevens et al, 2007;Do et al, 2010;Maloney et al, 2010;Quadrana et al, 2013Quadrana et al, , 2014. They have revealed the critical importance of cell wall invertase and fruit yield (Fridman et al, 2004;Ruan et al, 2012) and have identified the genomic regions underlying vitamin content in fruit (Stevens et al, 2007;Fitzpatrick et al, 2013;Quadrana et al, 2013Quadrana et al, , 2014.…”