Identification of transcription factors regulating plant secondary metabolism through the integration of functional genomics and metabolomics

植物产生非常广泛的代谢物，这些代谢物对于植物细胞的生长不是必需的，但是被植物用于提供针对胁迫和病原体的保护，以吸引传粉者和传播媒介，以及作为发育的信号。这些通常被称为“次级代谢产物”，但更普遍地被称为植物“天然产物”。天然产品最近被认为是饮食的重要组成部分，可以预防心血管疾病，某些癌症和与年龄相关的退行性疾病。它们也是美容产品和疾病自然疗法的重要组成部分。植物通常将其天然产物积累到相对较低的水平，因此人们对育种或工程植物产生更高水平的兴趣。研究表明，增加次生代谢产物积累的最有效方法是增加调节生物合成途径活性的基因的活性，这些途径产生不同的天然产物。这种类型的调节基因编码称为转录因子的蛋白质。使用这种策略来开发积累显著更高水平的重要天然产物的植物的最大瓶颈是，还没有在分子水平上鉴定出许多调节次级代谢的转录因子。该建议旨在从模式植物拟南芥中鉴定控制次生代谢不同分支的转录因子。这将通过研究一组选定的基因来实现，这些基因可能编码转录因子（从初步证据中确定为可能的调节因子）。38个这样的基因中的每一个的活性都将增加，对代谢的影响将通过物理化学技术来测量，这些技术可以确定植物提取物中小有机分子（代谢物）水平的主要差异。很可能几个选定的基因编码具有相同调节活性的转录因子。这些基因将被识别，因为增加它们的活性将导致植物中代谢物的完全相同的变化。目的是鉴定具有相同调节活性的所有蛋白质，然后从每个不同的活性组中选择一个实例，以确定该活性丧失对代谢物水平的影响。这将使用灵敏的分离技术结合质谱定量鉴定来完成。代谢指纹图谱和代谢产物谱的组合应该使我们能够识别新的基因编码的转录因子控制次生代谢，可用作工程天然产物积累的工具。该项目还将为植物代谢物谱的大范围鉴定和比较方法的开发做出重要贡献。

Plants produce a very broad array of metabolites, which are not essential for growth of plant cells, but which are used by plants to provide protection against stress and pathogens, to attact pollinators and dispersal agents and as signals for development. These are often referred to as 'secondary metabolites' but are known more generally as plant 'natural products'. Natural products have recently become recognised as important components of the diet, offering protection against cardiovascular diseases, certain cancers and age-related degenerative diseases. They are also important components of beauty products and natural remedies for diseases. Plants often accumulate their natural products to relatively low levels, so there is a lot of interest in breeding or engineering plants that produce higher levels. It has been shown that the most effective way to increase the accumulation of secondary metabolites is to increase the activity of genes that regulate the activity of the biosynthetic pathways that make different natural products. Regulatory genes of this type encode proteins called transcription factors. The biggest bottleneck in using this strategy to develop plants that accumulate significantly higher levels of important natural products is that not many transcription factors regulating secondary metabolism have yet been identified at the molecular level. This proposal aims to identify transcription factors from the model plant, Arabidopsis thaliana, that control different branches of secondary metabolism. This will be achieved by studying a selected group of genes that probably encode transcription factors (identified from preliminary evidence as likely regulators). The activity of each of 38 such genes will be increased and the effects on metabolism will be measured by physico-chemical techniques that can identify major differences in the levels of small organic molecules (metabolites) within plant extracts. It is likely that several of the selected genes encode transcription factors that have the same regulatory activity. These genes will be identified because increasing their activity will result in exactly the same changes in metabolites in plants. The aim will be to identify all the proteins that have the same regulatory activities and then to choose one example from each distinct activity group to establish the effects of loss of that activity on metabolite levels. This will be done using sensitive techniques of separation coupled with quantitative identification by mass spectrometry. The combination of metabolic fingerprinting and metabolite profiling should allow us to identify new genes encoding transcription factors controlling secondary metabolism that can be used as tools for engineering natural product accumulation. This project will also make an important contribution to method development for broad range identification and comparison of metabolite profiles in plants.