Transcription factor CsHSFB2c suppresses CsTS1 and CsGS1 expression to reduce theanine biosynthesis in tea plants under heat stress

Heat stress reduces theanine content in tea plants, but the underlying molecular mechanism remains unclear. In this study, a temperature gradient treatment (20℃, 25℃, 30℃, and 35℃) was performed to unveil the effect of heat stress on biosynthesis and accumulation of theanine. We found that heat stress induced metabolic changes, characterized by decreased theanine content and increased catechin levels. In addition, heat stress up-regulated the expression of the class B heat shock transcription factor gene CsHSFB2c, while significantly suppressing the transcription of key theanine biosynthetic genes CsTS1 and CsGS1. Functional studies showed that silencing CsHSFB2c increased theanine content, while its overexpression significantly decreased theanine levels. Consistent with these changes, silencing CsHSFB2c upregulated the expression of CsTS1 and CsGS1, while overexpression of CsHSFB2c downregulated their expression. Yeast one-hybrid (Y1H) and dual-luciferase reporter gene (Dual-LUC) assays showed that CsHSFB2c directly binds to the promoters of CsTS1 and CsGS1 and inhibits their expression. These results demonstrate that CsHSFB2c mediates heat-induced suppression of theanine biosynthesis by directly inhibiting the expression of CsTS1 and CsGS1. This study provides a theoretical basis for improving the heat resistance and quality of tea plants via molecular breeding.