CsGDH2.1 negatively regulates theanine accumulation in late-spring tea plants (Camellia sinensis var. sinensis)

Theanine, a unique and the most abundant non-proteinogenic amino acid in tea plants, endows tea infusion with the umami taste and anti-stress effects. Its content in tea correlates highly with green tea quality. Theanine content in new shoots of tea plants is high in mid-spring and greatly decreases in late spring. However, how the decrease is regulated is largely unknown. In a genetic screening, we observed that a yeast mutant, glutamate dehydrolase 2 (gdh2), was hypersensitive to 40 mM theanine and accumulated more theanine. This result implied a role of CsGDH2s in theanine accumulation in tea plants. Therefore, we identified the two homologs of GDH2, CsGDH2.1 and CsGDH2.2, in tea plants. Yeast complementation assay showed that the expression of CsGDH2.1 in yeast gdh2 mutant rescued the theanine hypersensitivity and hyperaccumulation of this mutant. Subcellular localization and tissue-specific expression showed CsGDH2.1 localized in the mitochondria and highly expressed in young tissues. Importantly, CsGDH2.1 expression was low in early spring, and increased significantly in late spring, in the new shoots of tea plants. These results all support the idea that CsGDH2.1 regulates theanine accumulation in the new shoots. Moreover, the in vitro enzyme assay showed that CsGDH2.1 had glutamate catabolic activity, and knockdown of CsGDH2.1 expression increased glutamate and theanine accumulation in the new shoots of tea plants. These findings suggested that CsGDH2.1-mediated glutamate catabolism negatively regulates theanine accumulation in the new shoots in late spring, and provides a functional gene for improving late-spring green tea quality.

Automated summary

Theanine, a non-proteinogenic amino acid abundant in tea plants, gives tea infusion its unique taste and anti-stress effects. The content of theanine in tea is closely correlated with its quality. However, the regulation of theanine decrease in late spring is still unknown. Through genetic screening, a yeast mutant, gdh2, was found to be hypersensitive to theanine and accumulated more of it, indicating a potential role of CsGDH2.1 in theanine accumulation in tea plants. CsGDH2.1 was found to be expressed in the mitochondria and highly expressed in young tissues of tea plants. CsGDH2.1 was also observed to regulate theanine accumulation in new shoots in late spring through glutamate catabolism.