Abstract：
Flavonols are widely synthesized throughout the plant kingdom, playing essential roles in plant physiology and providing unique health benefits for humans. Their glycosylation plays significant role in improving their stability and solubility, thus their accumulation and function. However, the genes encoding the enzymes catalyze this glycosylation remain largely unknown in apple. This study utilized a combination of methods to identify genes encoding such enzymes. Initially, candidate genes were selected based on their potential to encode UDP-dependent glycosyltransferases (UGTs) and their expression patterns in response to light induction. Subsequently, through testing the in vitro enzyme activity of the proteins produced in Escherichia coli cells, four candidates were confirmed to encode a flavonol 3-O-galactosyltransferase (UGT78T6), flavonol 3-O-glucosyltransferase (UGT78S1), flavonol 3-O-xylosyltransferase/arabinosyltransferase (UGT78T5), and flavonol 3-O-rhamnosyltransferase (UGT76AE22), respectively. Further validation of these genes\' functions was conducted by modulating their expression levels in stably transformed apple plants. As anticipated, a positive correlation was observed between the expression levels of these genes and the content of specific flavonol glycosides corresponding to each gene. Moreover, overexpression of a flavonol synthase gene, MdFLS, resulted in increased flavonol glycoside content in apple roots and leaves. These findings provide valuable insights for breeding programs aimed at enriching apple flesh with flavonols and for identifying flavonol 3-O-glycosyltransferases of other plant species.
摘要：
黄酮醇在整个植物界广泛合成，在植物生理学中发挥重要作用，为人类提供独特的健康益处。它们的糖基化在提高其稳定性和溶解性方面发挥着重要作用，因此，它们的积累和功能。然而,在苹果中，编码催化这种糖基化的酶的基因仍然是未知的。本研究利用多种方法的组合来鉴定编码此类酶的基因。最初，选择候选基因是基于它们编码UDP依赖性糖基转移酶（UGT）的潜力及其响应于光诱导的表达模式。随后，通过测试大肠杆菌细胞产生的蛋白质的体外酶活性，四个候选被证实编码黄酮醇3-O-半乳糖基转移酶（UGT78T6），黄酮醇3-O-葡萄糖基转移酶（UGT78S1），黄酮醇3-O-木糖基转移酶/阿拉伯糖基转移酶（UGT78T5），和黄酮醇3-O-鼠李糖基转移酶（UGT76AE22），分别。通过调节这些基因在稳定转化的苹果植株中的表达水平来进一步验证这些基因的功能。如预期,这些基因的表达水平与每个基因对应的特定黄酮醇苷含量呈正相关。此外，黄酮醇合酶基因的过表达，MdFLS,导致苹果根和叶中黄酮醇苷含量增加。这些发现为旨在使苹果肉富含黄酮醇和鉴定其他植物物种的黄酮醇3-O-糖基转移酶的育种计划提供了有价值的见解。
