Geraniol is an important contributor to the pleasant floral scent of tea products and one of the most abundant aroma compounds in tea plants; however, its biosynthesis and physiological function in response to stress in tea plants remain unclear. Here, we studied eight terpene synthases with expression levels that were correlated with geraniol accumulation in different tissues of tea plants. The proteins encoded by the full-length terpene synthase ( CsTPS1) and its alternative splicing isoform ( CsTPS1- AS) could catalyze the formation of geraniol when GPP was used as a substrate in vitro, whereas the expression of CsTPS1- AS was only significantly induced by Colletotrichum gloeosporioides and Neopestalotiopsis sp. infection. Silencing of CsTPS1 and CsTPS1- AS resulted in a significant decrease in the geraniol content in tea plants. The geraniol content and antifungal activity of tea plants were compared when CsTPS1 and CsTPS1- AS were silenced. Down-regulation of the expression of CsTPS1- AS reduced the accumulation of geraniol, and the silenced tea plants exhibited greater susceptibility to pathogen infection than control plants. However, the geraniol content and pathogen resistance of CsTPS1-silenced plants and control plants did not significantly differ. Further analysis showed that silencing of CsTPS1- AS led to a decrease in the expression of the defense-related genes PR1 and PR2 and the expression of SA pathway-related genes in tea plants, which increased the susceptibility of tea plants to pathogenic fungal infections . Both in vitro and in vivo results indicated that CsTPS1 is involved in the regulation of geraniol formation and plant defense via alternative splicing in tea plants. The results of this study provide new insights into geraniol biosynthesis and highlight the role of monoterpene synthases in modulating plant disease resistance via alternative splicing.

The biotechnological production of glycosides is an economically competitive manufacturing alternative to classical chemical synthesis. Through continuous production improvement, glycosides can now be used in low-cost products by various industries. However, many production systems still suffer from low yields. Directed evolution, coupled with a suitable screening method, can tackle this challenge. We generated glycosyltransferase mutants through error-prone-PCR and screened the library using a small-scale whole-cell biotransformation system. The screening of only 176 colonies yielded three putative candidates. Detailed investigations revealed that the reason for the increase in product titer was mainly due to different expression effects of the mutagenized genes rather than improved enzyme kinetics. In total, a 60-fold increase in product formation was achieved. Therefore, in addition to the quality of the mutant library, an efficient and stable expression system is crucial to achieve high concentrations of active enzyme and product, as formation of inclusion bodies and other inactive forms of the biocatalyst reduces productivity.