Gibberellin promotes theanine synthesis by relieving the inhibition of CsWRKY71 on CsTSI in tea plant (Camellia sinensis)

Theanine is a crucial indicator of tea quality, its significance is closely tied to the economic value of tea. There were many reports on the regulation mechanism of theanine synthesis and accumulation, but the mechanism of gibberellin regulates theanine synthesis in tea plants is poorly understood. Previous studies have shown that the content of theanine experiences significant changed in the growth stages of tea shoots, displayed a strong correlation with gibberellin. This study confirmed that gibberellin significantly promoted the expression of the major gene of theanine synthesis, known as CsTSI. Additionally, the study identified CsWRKY71, a transcription factor mediated the regulation of gibberellin on theanine synthesis in tea plants. CsWRKY71 was localized in the nucleus and had a typical WRKY domain. It was a member of class IIC sub- and its expression was significantly suppressed following exogenous GA3 treatment. Further assay such as EMSA, dual luciferase and asODN interfering demonstrated that CsWRKY71 significantly interacted with the promoter of CsTSI, which inhibited theanine synthesis by binding to the cis-acting element (C/T)TGAC(T/C) of CsTSI promoter. Overall, the addition of exogenous gibberellin alleviated the inhibition of CsTSI by down-regulating the expression of CsWRKY71, ultimately facilitated the rapid biosynthesis of theanine. This study elucidated the molecular mechanism of CsWRKY71 mediated gibberellin regulation of theanine synthesis in tea plant. The findings not only enhance our understanding of the regulatory processes involved in theanine synthesis in tea plants, but also provide important references for maintaining the characteristics of high theanine in tea plant.