REGULATION OF AMINO ACID BIOSYNTHESIS GENES IN PLANTS

植物氨基酸生物合成基因的调控

基本信息

批准号：
6695673
负责人：
Gloria CORUZZI
金额：
$ 2.43万
依托单位：
NEW YORK UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1983
资助国家：
美国
起止时间：
1983-12-01 至 2004-02-29
项目状态：
已结题

项目摘要

The overall goal of our research is to gain a molecular understanding of the regulatory processes that control the assimilation of inorganic nitrogen in plants. This process plays a central role in the regulation of plant growth and development. Using molecular-genetic approaches in Arabidopsis, we identified isoenzymes of GS, GOGAT and GDH that control the assimilation of inorganic-N into Glu/Gln, key amino acids used to transport nitrogen within and between cells. Our studies indicate that expression of these genes is regulated by the metabolic status of the plant. For example, transcriptional induction of GS by light, can be mimicked by sucrose in the absence of light. Moreover, sucrose induction of GS expression can be antagonized by amino acids, which results in repression of GS activity. This led us to hypothesize that plants have a mechanism to sense internal levels of amino acids. This would allow a plant to turn off assimilation of inorganic-N when internal levels of amino acid are high. Testing this hypothesis, defining it mechanistically, and identifying components thereof, is the focus of this renewal. Towards this goal, we have begun to characterize amino acid sensing/signaling components in Arabidopsis using molecular-genetic, cell biological, and biochemical approaches. Our reverse genetic studies are driven by the hypothesis that amino acid sensing is primitive and conserved in evolution. In support of this, the repression of GS expression by amino acids in plants is mechanistically reminiscent of the Ntr system in E. coli. Moreover, we identified a plant homologue of an Ntr component, PII, and showed using PII transgenic plants that PII appears to play a role in C:N sensing in chloroplasts an in GS regulation, as it does in Ntr. The amino acid products of N-assimilation are exported from chloroplasts and transported to other cells, and we have evidence that Glu, the prinicple intermediate, may serve as an extracellular signal . In support of this, we identified putative sensors of extracellular Glu, plant homologues of animal glutamate receptors (iGluRs). We showed plant GluRs function as ligand-gated ion channels, and studies of GLR transgenic plants indicate they may play a role in light signal transduction, reminiscent of their counterparts in the retina and brain. These findings suggest iGluRs are derived from a primitive amino acid signaling mechanism that existed before plants and animals diverged. We propose to exploit this evolutionary conservation and test whether Arabidopsis (or GLR mutants we isolate) can be used in a bioassay for drugs to treat GluR-related diseases in humans. We will also use forward genetic approaches to isolate amino acid sensing/signaling mutants in Arabidopsis which may identify components of these evolutionarily conserved amino acid signaling pathways or novel pathways.

我们研究的总体目标是从分子角度了解控制植物中无机氮同化的调节过程。这个过程在植物生长和发育的调节中起着核心作用。利用拟南芥中的分子遗传学方法，我们鉴定了 GS、GOGAT 和 GDH 的同工酶，它们控制无机氮同化为 Glu/Gln（用于在细胞内和细胞间运输氮的关键氨基酸）。我们的研究表明这些基因的表达受到植物代谢状态的调节。例如，在没有光的情况下，蔗糖可以模拟光对 GS 的转录诱导。此外，蔗糖诱导的 GS 表达可以被氨基酸拮抗，从而抑制 GS 活性。这使我们推测植物具有感知内部氨基酸水平的机制。当内部氨基酸水平较高时，这将使植物关闭无机氮的同化。检验这一假设、机械地定义它并确定其组成部分是本次更新的重点。为了实现这一目标，我们已经开始使用分子遗传学、细胞生物学和生化方法来表征拟南芥中的氨基酸传感/信号成分。我们的反向遗传学研究是基于氨基酸传感在进化中是原始且保守的假设。为了支持这一点，植物中氨基酸对 GS 表达的抑制在机制上让人想起大肠杆菌中的 Ntr 系统。此外，我们鉴定了 Ntr 成分 PII 的植物同源物，并使用 PII 转基因植物表明，PII 似乎在叶绿体中的 C:N 感应和 GS 调节中发挥作用，就像在 Ntr 中一样。 N同化的氨基酸产物从叶绿体输出并转运到其他细胞，并且我们有证据表明主要中间体Glu可能充当细胞外信号。为了支持这一点，我们确定了细胞外谷氨酸的假定传感器，即动物谷氨酸受体（iGluR）的植物同系物。我们展示了植物 GluR 作为配体门控离子通道的功能，对 GLR 转基因植物的研究表明它们可能在光信号转导中发挥作用，让人想起它们在视网膜和大脑中的对应物。这些发现表明 iGluR 源自植物和动物分化之前就存在的原始氨基酸信号传导机制。我们建议利用这种进化保守性并测试拟南芥（或我们分离的 GLR 突变体）是否可用于治疗人类 GluR 相关疾病的药物生物测定。我们还将使用正向遗传方法来分离拟南芥中的氨基酸传感/信号传导突变体，这可能会识别这些进化上保守的氨基酸信号传导途径或新途径的成分。