近年来, UGTs基因在植物响应非生物胁迫中的作用逐渐被揭示。基于转录组数据, 筛选出两个在低温胁迫下显著上调, 且与茶树(Camellia sinensis)花青苷合成相关的糖基转移酶基因CsUGT73B4(CSS0039619)和CsUGT85K5(CSS0047548)。为探究两个基因在响应低温胁迫下调控茶树花青苷代谢的分子机制, 我们克隆了CsUGT73B4和CsUGT85K5并进行了生物信息学分析, 通过拟南芥过表达和茶树瞬时沉默实验验证其功能。结果显示: 低温胁迫下CsUGT73B4/CsUGT85K5基因的表达与花青苷积累呈正相关; 过表达拟南芥中, 花青苷含量、抗氧化酶活性(SOD、POD、CAT)显著提高, 而丙二醛和相对电导率显著下降; 在茶树中瞬时沉默CsUGT73B4或CsUGT85K5后, 花青苷含量及其合成相关基因的表达水平均显著下调。本研究初步探讨了茶树CsUGT73B4和CsUGT85K5基因的功能, 揭示了其通过调控花青苷代谢提高茶树抗寒性的分子机制, 为茶树抗寒分子育种提供了相关候选基因。

INTRODUCTION:  Glycosyltransferases are key enzymes involved in various biological processes in plants. To date, glycosyltransferases have been classified into 138 families, among which the glycosyltransferase family I is the largest. This family primarilyuses UDP-glucose as the glycosyl donor and is referred to as the UGT family. In recent years, the role of UGT genes in plant responses to abiotic stresses has gradually been revealed. However, currently, only a few UGT genes have been clearly identified as being involved in anthocyanin biosynthesis.

RATIONALE: To explore the mechanisms of anthocyanin and its glycosylation-related genes in tea plants under low-temperature stress, this study used FudingDabai Tea as the experimental material. Based on the transcriptome sequencing data, two genes, CsUGT73B4 (CSS0039619) and CsUGT85K5 (CSS0047548), which significantly upregulate anthocyanin expression, were identified. We further investigated the changes in these genes under low-temperature stress and explored the relationship between CsUGT73B4 and CsUGT85K5 genes and anthocyanin metabolism under low-temperature stress. Additionally, the functions of these two genes were verified through heterologous overexpression in Arabidopsis thaliana and gene silencing experiments.

RESULTS: Under low-temperature stress, the expression of CsUGT73B4 and CsUGT85K5 genes was positively correlated with anthocyanin accumulation. In Arabidopsis thaliana plants overexpressing these genes, anthocyanin content and antioxidant enzyme activities (SOD, POD, CAT) were significantly increased, while malondialdehyde (MDA) content and relative conductivity were reduced. Silencing of CsUGT85K5 led to feedback inhibition of CsUGT73B4 expression, revealing that these two genes act in concert to regulate the anthocyanin metabolic pathway.

CONCLUSION: In summary, the CsUGT73B4 and CsUGT85K5 genes are involved in the response to low-temperature stress, regulate anthocyanin metabolism, and may enhance the cold tolerance of tea plants.

The effect of low-temperature stress on anthocyanin in WT、OE-CsUGT73B4 and OE-CsUGT85K5

(A) Observation of anthocyanin accumulation phenotype in Arabidopsis, the apical meristem position was marked with a white thread (The white line bar=1mm); (B) Effect of low-temperature stress on anthocyanin content in WT and OE-CsUGT73B4 Arabidopsis; (C) Effect of low-temperature stress on anthocyanin content in WT and OE-CsUGT85K5 Arabidopsis.  *P<0.05, **P<0.01, ***P<0.001.