A tonoplast Glu/Asp/GABA exchanger that affects tomato fruit amino acid composition.
A tonoplast Glu/Asp/GABA exchanger that affects tomato fruit amino acid composition.
复制标题
DOI：
10.1111/tpj.12766
复制
发表时间：
2015-03
期刊：
影响因子：
--
通讯作者：
Sweetlove LJ
中科院分区：
文献类型：
--
作者：
Snowden CJ;Thomas B;Baxter CJ;Smith JA;Sweetlove LJ
番茄果实中酸性物质的积累是影响果实风味和营养品质的重要因素。氨基酸Asp和Glu在成熟过程中积累到高浓度，而γ-氨基丁酸（GABA）显示出近似化学计量的下降。考虑到GABA可以被分解代谢形成Glu和随后的Asp，以及果实在成熟过程中保持渗透平衡的要求，我们假设存在一种液泡膜转运蛋白，该转运蛋白从液泡中出口GABA以换取Asp或Glu的进口。我们在这里表明，番茄液泡膜具有这样的运输特性：谷氨酸通过孤立的液泡膜的运输是反式刺激的反交换模式的GABA，Glu和Asp。我们使用富含液泡膜组分的定量蛋白质组学鉴定了SlCAT 9作为这种交换剂的候选蛋白。SlCAT 9-YFP融合体在烟草中的瞬时表达证实了液泡膜定位。通过在转基因番茄植物中过表达SlCAT 9来检查蛋白质的功能。从转基因植物中分离的液泡膜囊泡仅在与Glu、Asp或GABA以反向交换模式测定时显示出比野生型（WT）更高的Glu和GABA转运速率。此外，转基因植物成熟果实中所有三种同源氨基酸的含量均显著增加。我们得出结论，SlCAT 9是一个液泡膜Glu/Asp/GABA交换器，强烈影响这些氨基酸在果实发育过程中的积累。番茄果实在成熟过程中的氨基酸组成发生重大变化，数量上最重要的变化是GABA的下降和Asp和Glu的化学计量增加。我们已经确定了一种液泡膜转运蛋白SlCAT 9，它促进GABA / Glu / Asp在果实液泡膜上的反向交换，并强烈影响成熟果实中这三种氨基酸的水平。
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 (GABA) shows an approximately stoichiometric decline. Given that GABA 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 GABA 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 trans‐stimulated in counterexchange mode by GABA, Glu and Asp. We identified SlCAT9 as a candidate protein for this exchanger using quantitative proteomics of a tonoplast‐enriched membrane fraction. Transient expression of a SlCAT9‐YFP fusion in tobacco confirmed a tonoplast localisation. The function of the protein was examined by overexpression of SlCAT9 in transgenic tomato plants. Tonoplast vesicles isolated from transgenic plants showed higher rates of Glu and GABA transport than wild‐type (WT) only when assayed in counterexchange mode with Glu, Asp, or GABA. Moreover, there were substantial increases in the content of all three cognate amino acids in ripe fruit from the transgenic plants. We conclude that SlCAT9 is a tonoplast Glu/Asp/GABA exchanger that strongly influences the accumulation of these amino acids during fruit development. Tomato fruit undergo major shifts in amino acid composition during ripening with the quantitatively most important changes being a decline in GABA and a stoichiometric increase in Asp and Glu. We have identified a tonoplast transporter, SlCAT9, that facilitates the counter‐exchange of GABA / Glu / Asp across the fruit vacuolar membrane and strongly influences the levels of the three amino acids in ripe fruit.