Methionine Biosynthesis in Higher Plants

高等植物中的蛋氨酸生物合成

• 批准号: 9728661
• 负责人: Thomas Leustek
• 金额: $ 30.63万
• 依托单位: Rutgers University New Brunswick
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-03-01 至 2002-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9728661&HistoricalAwards=false
• 关键词: Methionine; Biosynthesis; Higher; Plants

9728215 Leustek The topic of this project is the developmental regulation of Met biosynthesis in higher plants, focusing primarily on the key enzyme cystathionine (-synthase (CS), and with some emphasis on threonine synthase (TS) and homoserine kinase (HK). The goal is to study how Met synthesis is regulated in cells and tissues during development in the model vascular plant Arabidopsis thaliana. Using a reporter gene method it was determined that expression from the CS promoter is not equal in all cells. In seedlings the CS promoter is highly active in meristems and the vascular tissues of roots and cotyledons. The activity is significantly lower in mesophyll cells. But in this cell type it is induced by treatment of seedlings with Thr & Lys (inhibitors of Met synthesis), and is repressed by Met-feeding. Promoter activity in the vascular tissues is unaffected by these treatments. In mature plants the CS promoter is active primarily in vascular tissue of leaves, the flower stalk, floral receptacle and anthers; and is inactive in siliques and developing seeds. Inhibition of CS expression with antisense-RNA causes a severe growth defect that can be partially restored by Met-feeding. The phenotype of these plants resembles that of A. thaliana with reduced levels of DNA methylation. The next phase of the project will center on further exploration of the reporter gene results. The possible role of substrate competition in regulating Met synthesis will be explored by studying CS and TS expression in parallel. Both will be analyzed by measurement of in situ mRNA and protein. The affect of Met and its metabolites on cell specific expression will be studied in two ways: through analysis of CS and TS in plants that have been treated with agents that perturb Met synthesis, and through analysis of A. thaliana mutant lines with elevated levels of Met or with lower levels of S-adenosylmethionine. The subcellular localization of the CS enzyme in plant cells will be explored and the kinetic propert ies of the recombinant CS enzyme will be analyzed in order to determine whether enzyme regulation plays any role regulating Met synthesis. Finally, the cDNA encoding HK will be cloned, the goal being to study the HK enzyme and to study how expression of HK is regulated. The amino acid methionine is of vital importance to all living organisms being intimately involved in every aspect of metabolism. Animals are unable to produce methionine and they must obtain it from their diet. Yet, most crop plants do not contain sufficient amounts of this key nutrient. One reason may be that plants control precisely the amount of methionine that they make. Understanding how this regulation takes place could lead to methods for improving the nutritional value of crops by increasing their methionine content.

小行星9728215 本项目的主题是高等植物Met生物合成的发育调控，主要集中在关键酶胱硫醚合成酶（CS），并重点关注苏氨酸合成酶（TS）和高丝氨酸激酶（HK）。 目的是研究在维管植物拟南芥（Arabidopsis thaliana）的发育过程中，细胞和组织中Met的合成是如何受到调控的。使用报告基因方法，确定CS启动子的表达在所有细胞中不相等。在幼苗中，CS启动子在根和子叶的分生组织和维管组织中高度活跃。叶肉细胞中的活性明显较低。但在这种细胞类型中，它是通过用Thr Lys（Met合成的抑制剂）处理幼苗来诱导的，并且被Met喂养所抑制。这些处理不影响维管组织中的启动子活性。在成熟植物中，CS启动子主要在叶、花茎、花托和花药的维管组织中有活性;而在长角果和发育中的种子中无活性。 用反义RNA抑制CS表达会导致严重的生长缺陷，可以通过Met喂养部分恢复。这些植物的表型与A.拟南芥的DNA甲基化水平降低。 该项目的下一阶段将集中在报告基因结果的进一步探索上。底物竞争在调节甲硫氨酸合成中的可能作用将通过平行研究CS和TS表达来探讨。两者都将通过原位mRNA和蛋白质的测量进行分析。Met及其代谢产物对细胞特异性表达的影响将通过两种方式进行研究：通过分析用干扰Met合成的试剂处理过的植物中的CS和TS，以及通过分析A.具有升高水平的Met或具有较低水平的S-腺苷甲硫氨酸的拟南芥突变系。将探索CS酶在植物细胞中的亚细胞定位，并分析重组CS酶的动力学性质，以确定酶调节是否在调节Met合成中起任何作用。 最后，将克隆编码HK的cDNA，目的是研究HK酶和研究HK的表达是如何调节的。 氨基酸蛋氨酸是至关重要的所有生物体密切参与代谢的各个方面。动物不能产生蛋氨酸，它们必须从饮食中获得。然而，大多数农作物并不含有足够量的这种关键营养素。其中一个原因可能是植物精确地控制它们制造的蛋氨酸的量。了解这种调节是如何发生的，可以通过增加蛋氨酸含量来提高作物的营养价值。