Defining the Tyrosine Biosynthetic Pathways in Plants

非技术总结：植物合成芳香族氨基酸酪氨酸（Tyr），这是人类饮食中的必需营养素，也是许多植物特异性化合物的关键前体。这些Tyr衍生的植物天然产物对植物生长、发育和环境响应具有深远的影响，并且在人类中也具有重要的营养和药理活性（例如，维生素E、吗啡）。尽管Tyr在植物和人类生物学中都很重要，但我们仍然不知道Tyr是如何在植物中合成的。因此，该项目将获得基础知识，以提高作物和药用植物的营养质量或药用价值，并有可能改善植物生长和胁迫反应，以实现粮食和生物质的可持续生产。一项名为“颜料艺术”的外联活动将利用来自芳香族氨基酸的有吸引力和有益的植物颜料（例如，甜菜色素）。虽然年幼的孩子会欣赏天然色素中美丽而复杂的颜色，但父母和年龄较大的孩子将有机会了解这些色素是如何由植物合成的。项目技术概述：越来越多的证据表明，许多植物通过中间体arogenate合成Tyr，因此与通常使用4-羟基苯丙酮酸作为中间体的微生物模型不同。然而，不同植物物种之间的Tyr生物合成途径似乎存在显着差异。该项目的目标是确定植物中Tyr的生物合成途径，并确定其区室化和关键调控步骤。拟议的研究将建立在PI最近分离的拟南芥和豆类基因编码arogenate和prephenate脱氢酶（ADH和PDH），酪氨酸途径的关键酶。拟南芥ADH对arogenate具有很强的底物特异性，定位于质体中，并被Tyr抑制。相比之下，豆类淀粉酶特异性地使用预苯酸盐，缺乏质体转运肽，并且对Tyr不敏感。本研究的主要假设是，拟南芥主要通过质体arogenate途径合成Tyr，而豆科植物通过arogenate和4-hydroxyphenylpyruvate合成Tyr，但其定位和调控机制尚不清楚。在这个项目中，动力学分析和本地化的研究，首先计划为新确定的豆科植物的PDHs（目标1）。在植物中的功能的PDHs和ADH将使用两个对比的植物模型，苜蓿（Aim 2）和拟南芥（Aim 3）进行调查和比较。将在生成的突变体和转基因体中进一步进行靶向代谢物分析和稳定同位素补料实验，以直接测量替代Tyr途径中碳流的分布。虽然初级代谢通常被认为是保守的跨植物物种，我们的初步数据表明，否则。该项目将直接测试这一假设，并解决Tyr生物合成（一种主要代谢途径）如何在不同植物中进化，以支持下游专门代谢中存在的巨大化学多样性。

Non-technical summary:Plants synthesize an aromatic amino acid tyrosine (Tyr), which is an essential nutrient in human diets and a key precursor of numerous plant specialized compounds. These Tyr-derived plant natural products have profound impacts on plant growth, development and environmental responses, and also have important nutritional and pharmacological activities in humans (e.g., vitamin E, morphine). Despite their importance in both plant and human biology, we still do not know how Tyr is synthesized in plants. Therefore, this project will gain fundamental knowledge to enhance the nutritional quality or pharmaceutical values of crops and medicinal plants, and can potentially improve plant growth and stress responses for sustainable production of food and biomass. An outreach event, "Pigment-Art", will take advantage of attractive and beneficial plant pigments derived from aromatic amino acids (e.g., betalains). While younger children will appreciate the beautiful and complex colors in natural pigments, parents and older children will have opportunities to learn about how these pigments are synthesized by plants. Technical Description of the Project:Accumulating evidence suggests that many plants synthesize Tyr via the intermediate arogenate and therefore differ from microbial models that usually use 4-hydroxyphenylpyruvate as an intermediate. However, significant variations appear to exist for the Tyr biosynthetic routes among different plant species. The goals of this project are to define the Tyr biosynthetic route(s) in plants and determine their compartmentalization and crucial regulatory step(s). The proposed research will build upon PI's recent isolation of Arabidopsis and legume genes encoding arogenate and prephenate dehydrogenases (ADH and PDH), key enzymes of the Tyr pathway. Arabidopsis ADHs have strong substrate specificity toward arogenate, are localized in the plastids and inhibited by Tyr. In contrast, legume dehydrogenases specifically use prephenate, lack a plastid transit peptide, and are insensitive to Tyr. The primary hypothesis of this study is that Arabidopsis mainly uses the plastidic arogenate pathway that is regulated by Tyr, whereas legumes synthesize Tyr via both arogenate and 4-hydroxyphenylpyruvate with unknown localization and regulatory mechanism. In this project, kinetic analysis and localization studies are first planned for the newly identified legume PDHs (Aim 1). In planta functions of PDHs and ADHs will be investigated and compared using two contrasting plant models, Medicago truncatula (Aim 2) and Arabidopsis thaliana (Aim 3). Targeted metabolite profiling and stable-isotope feeding experiments will be further conducted in the generated mutants and transgenics to directly measure the distribution of carbon flow among the alternative Tyr pathways. Although primary metabolism is generally assumed to be conserved across plant species, our preliminary data indicate otherwise. This project will directly test this assumption and address how Tyr biosynthesis, a primary metabolic pathway, has evolved in different plants to support the enormous chemical diversity that exists in the downstream specialized metabolism.