Vacuolar Sequestration of Anthocyanin in Maize

9603927种沃尔伯特开花植物合成了令人难以置信的多样化的“有毒”天然产物。PIs在医学上使用了许多这样的化合物，例如抗癌药物紫杉醇和秋水仙碱，心脏药物洋地黄，甚至广泛使用的止痛药阿司匹林和奎宁都是基于植物天然产品。在植物内部，这些化合物和许多额外的色素和次生代谢物阻止食草动物和病原体，因此是植物在自然界中生存能力的一部分。然而，令人惊讶的是，人们对植物如何在不损害自身细胞的情况下合成“有毒”分子却知之甚少。生物合成发生在植物细胞的细胞质中，但有毒的代谢产物储存在植物的液泡中。花青素生物合成的生物化学和分子生物学是高等植物中最被理解的次生代谢产物途径。花青素是在大多数花和许多水果中发现的红蓝紫三色色素，它们的颜色使它们成为阐明液泡隔离机制的有用“标记物”。在玉米中，Bronze2基因编码花青素生物合成的最后一个遗传步骤。这种酶是一种谷胱甘肽S转移酶(GST)，它为细胞质花青素增加了一个化学“标签”，促进了进入植物液泡的过程。这是植物将大量次生代谢物运输到液泡中的第一个已建立的机制。在这项拟议的工作中，研究人员将确定Bronze2酶的分布，确定它是否与其他GST络合，并确定该酶在体内除花青素外是否还有其他底物。Bz2和其他玉米GST中的突变将被用来定义哪些酶发挥独特的作用，哪些具有重叠的作用。对瞬时谷胱甘肽转化的花青素色素的结构研究将确定谷胱甘肽标记的位置(S)，并确定液泡中随后的花色苷修饰是否发生在细胞质中标记谷胱甘肽的相同位置。了解花青素从细胞质向液泡转移的关键步骤将为了解植物合成和储存广泛的有毒但非常有用的代谢物的能力提供新的见解。

9603927 Walbot Flowering plants synthesize an incredible diversity of "toxic" natural products. The PIs use many of these compounds in medicine, for example, the anti-cancer drugs taxol and colchicine, the heart medicine digitalis, and even the widely used pain relievers of aspirin and quinine are based on plant natural products. Within the plant, theses compounds and many additional pigments and secondary metabolites deter herbivores and pathogens, and are thus part of the plant's ability to survive in the natural world. Surprisingly little is known, however, about how plants can synthesize "toxic" molecules and yet avoid damaging their own cells. Biosynthesis occurs in the plant cell cytoplasm, but the toxic metabolites are stored in the plant vacuole. The biochemistry and molecular biology of anthocyanin pigment biosynthesis is the best understood secondary metabolite pathway in higher plants. Anthocyanins are the red-blue-purple pigments found in most flowers and many fruits, and their color makes them a useful "marker" for elucidating the mechanisms of vacuolar sequestration. In maize, the Bronze2 gene encodes that last genetically defined step in anthocyanin biosynthesis. This enzyme is a glutathione S-transferase (GST) that adds a chemical "tag" to cytoplasmic anthocyanin that facilitates entry into the plant vacuole. This is the first established mechanism by which plants can transport large secondary metabolites into the vacuole. In the proposed work, the investigators will define the distribution of the Bronze2 enzyme, determine if it complexes with other GSTs, and determine if this enzyme has additional in vivo substrates besides anthocyanin. Mutants in Bz2 and other maize GSTs will be used to define which enzymes perform unique roles and which have overlapping roles. Structural studies of the transiently glutathionationated anthocyanin pigment will define the site(s) of glutathione tagging and determine if subsequent anthocyanin modification in the vacuole occurs at th e same sites tagged with glutathione in the cytoplasm. Understanding the key steps of anthocyanin transfer from cytoplasm to vacuole will provide new insights into the capacity of plants to synthesize and store a wide range of toxic but very useful metabolites.