A tonoplast Glu/Asp/<scp>GABA</scp> exchanger that affects tomato fruit amino acid composition

<jats:title>Summary</jats:title><jats:p>Vacuolar accumulation of acidic metabolites is an important aspect of tomato fruit flavour and nutritional quality. The amino acids Asp and Glu accumulate to high concentrations during ripening, while γ‐aminobutyrate (<jats:styled-content style="fixed-case">GABA</jats:styled-content>) shows an approximately stoichiometric decline. Given that <jats:styled-content style="fixed-case">GABA</jats:styled-content> can be catabolised to form Glu and subsequently Asp, and the requirement for the fruit to maintain osmotic homeostasis during ripening, we hypothesised the existence of a tonoplast transporter that exports <jats:styled-content style="fixed-case">GABA</jats:styled-content> from the vacuole in exchange for import of either Asp or Glu. We show here that the tomato vacuolar membrane possesses such a transport property: transport of Glu across isolated tonoplast vesicle membranes was <jats:italic>trans</jats:italic>‐stimulated in counterexchange mode by <jats:styled-content style="fixed-case">GABA</jats:styled-content>, Glu and Asp. We identified Sl<jats:styled-content style="fixed-case">CAT</jats:styled-content>9 as a candidate protein for this exchanger using quantitative proteomics of a tonoplast‐enriched membrane fraction. Transient expression of a Sl<jats:styled-content style="fixed-case">CAT</jats:styled-content>9‐<jats:styled-content style="fixed-case">YFP</jats:styled-content> fusion in tobacco confirmed a tonoplast localisation. The function of the protein was examined by overexpression of Sl<jats:styled-content style="fixed-case">CAT</jats:styled-content>9 in transgenic tomato plants. Tonoplast vesicles isolated from transgenic plants showed higher rates of Glu and <jats:styled-content style="fixed-case">GABA</jats:styled-content> transport than wild‐type (<jats:styled-content style="fixed-case">WT</jats:styled-content>) only when assayed in counterexchange mode with Glu, Asp, or <jats:styled-content style="fixed-case">GABA</jats:styled-content>. Moreover, there were substantial increases in the content of all three cognate amino acids in ripe fruit from the transgenic plants. We conclude that Sl<jats:styled-content style="fixed-case">CAT</jats:styled-content>9 is a tonoplast Glu/Asp/<jats:styled-content style="fixed-case">GABA</jats:styled-content> exchanger that strongly influences the accumulation of these amino acids during fruit development.</jats:p>