Unravelling the transcription factor network controlling the biosynthesis of tryptophan-derived antimicrobial compounds in Arabidopsis roots and shoots

作为一种有效的防御策略，植物产生具有抗微生物特性的次生代谢产物来击败入侵的微生物。在模式植物拟南芥中，Camalexin和吲哚硫代葡萄糖苷(IGs)都是从主要代谢物色氨酸(Trp)中衍生出来的，并已被证明能抑制各种植物病原菌的生长。导致色氨酸衍生二次化合物的复杂生物合成途径在转录水平上是高度协调的，因此为研究转录调控网络提供了一个很好的模型系统。一些转录因子(TF)已被确定控制植物地上部分的代谢途径基因。初步结果支持这样一种观点，即在根中工作的转铁蛋白不同于叶，并且尚未完全解决。该项目旨在通过根和叶组织的比较，确定色氨酸衍生次生代谢的完整和特定分支的假定转录主要调节因子。为了确定根中新的调控因子，将应用根原生质体高通量筛选系统(Protoplast Trans Activation，PTA)，对1500多个拟南芥TF进行分析。候选转录因子将通过诱导获得和功能丧失的方法进行功能表征，包括TF对靶基因调控的影响、使用染色质免疫沉淀(ChIP)的直接启动子结合、调节启动子顺式元件、代谢物变化和病原体防御。结合已有的知识和新发现的转录因子的数据，我们的目标是揭示在植物防御中这一关键代谢途径的协调转录调控基础上的分层调控因子网络。此外，对这些调控电路的洞察将支持进一步改造抗病作物的尝试。

As an efficient defense strategy, plants produce secondary metabolites with antimicrobial properties to defeat invading microorganisms. In the model plant Arabidopsis, camalexin and indole-glucosinolates (IGs) are both derived from the primary metabolite tryptophan (Trp) and have been demonstrated to inhibit growth of various plant pathogens. The complex biosynthetic pathway leading to Trp-derived secondary compounds is highly coordinated on transcriptional level and therefore, provides an excellent model system for studying transcriptional regulatory networks. Several transcription factors (TFs) have been identified to control the metabolic pathway genes in the above-ground part of the plant. Preliminary results support the notion that the TFs operating in roots differ from leaves and are yet completely unresolved. This project aims to identify putative transcriptional master regulators of the complete and of specific branches of Trp-derived secondary metabolism comparing root and leaf tissues. To identify novel regulators in roots, a high-throughput screening system in root protoplasts (Protoplast Trans Activation, PTA) will be applied enabling analysis of more than 1500 Arabidopsis TFs. Candidate TFs will be functionally characterized by inducible gain- and loss-of-function approaches with respect to the TF impact on target gene regulation, direct promoter binding using Chromatin Immunoprecipitation (ChIP), regulatory promoter cis-elements, altered metabolite profile and pathogen defense. Combining pre-existing knowledge with data on the newly identified TFs, we aim to disclose the hierarchical regulatory TF networks underlying co-ordinated transcriptional regulation of this crucial metabolic pathway in plant defense. Moreover, insights in these regulatory circuits will support further attempts to engineer disease resistant crop plants.