A Biochemical Genetic Analysis of Inositol Metabolism in Subcellular Organelles

约翰逊肌醇是一种六碳环醇，对真核细胞的生长、分化和通讯至关重要。肌醇磷酸酯为多种激素、生长因子和神经递质传递信号。因此，Insitol的代谢在许多农业和临床学科中是一个重要的问题。尽管大量的生理和生化数据证明了肌醇对高等植物的重要性，但对调节其代谢的分子遗传机制知之甚少。本研究的目的是了解植物生长发育过程中控制肌醇代谢的遗传机制。研究者对肌醇生物合成的发育研究表明，关键的生物合成酶肌肌醇-1-磷酸合成酶（Ml-1-P合成酶）存在，其中一种形式定位于菜豆和拟南芥的叶绿体类囊体。鉴于这一发现，有必要对目前磷脂酰肌醇（PI）生物合成的两种途径模型进行研究。根据该模型，PI（磷脂苷的前体）在内质网（ER）中合成，然后转运到其他亚细胞细胞器，如叶绿体。然而，研究人员最近的发现（在叶绿体中合成肌醇）提出了PI和磷脂苷也在这些细胞器中合成的可能性。可以想象，肌醇和/或磷酸肌醇信号转导途径是叶绿体和细胞核之间通信系统的一部分（植物中假设的叶绿体到细胞核信号转导途径）。研究者将使用酶分析、western分析、免疫细胞化学分析、原位杂交、磷脂研究和分子分析来(1)将肌醇生物合成位点与PI生物合成位点联系起来；(2)分离肌醇代谢缺陷突变体。该项目有望为肌醇在叶绿体和其他亚细胞细胞器中合成肌醇的功能提供直接证据。这些发现将直接适用于对重要的植物磷酸肌苷信号转导途径的全面了解（仍然缺乏）。本课题研究植物中肌醇的合成。虽然我们对动物体内肌醇磷酸信号传导已经了解很多，但对植物体内肌醇磷酸信号传导的相关问题还没有很好的研究。这项工作的总体长期目标是了解肌醇在亚细胞细胞器生物学中的作用。研究者在先前的研究中表明，关键的生物合成酶肌醇-1-磷酸合成酶具有时间和空间控制的同型；这一意想不到的发现是新颖的。该项目将包括对肌醇和磷酸肌醇合成位点的详细研究。结合免疫细胞化学分析，原位杂交研究和酶活性测量将使用菜豆和拟南芥进行。这项工作将有助于发展对植物细胞信号传导的理解，以及对重要代谢途径的调节。

9724117 Johnson Inositol, a six-carbon cyclitol, is essential for growth, differentiation, and communication of eukaryotic cells. Inositol phosphates convey signals for a wide variety of hormones, growth factors and neurotransmitters. Insitol's metabolism, therefore, is an important concern in many agricultural and clinical disciplines. Although extensive physiological and biochemical data document the importance of inositol to higher plants, less is known of the molecular genetic mechanisms regulating its metabolism. The goal of this research is to understand the genetic mechanisms controlling inositol metabolism during plant growth and development. The investigator's developmental studies of inositol biosynthesis have shown, that isoforms of the pivotal biosynthetic enzyme, myo-inositol-1- phosphate synthase (Ml-1-P synthase), exist and that one form is localized to chloroplast thylakoids in Phaseolus vulgaris and Arabidopsis thaliana. Given this finding, it is imperative that the current two pathway model for phosphatidylinositol (PI) biosynthesis in plans be examined. According to this model, PI(precursor for phosphinositdes) is synthesized in the endoplasmic reticulum (ER) and then transported into other subcellular organelles such as the chloroplasts. The investigator's recent finding (the synthesis of inositol in chloroplasts), however, raises the possibility that PI and the phosphinositides are also being synthesized in these organelles. It is conceivable that inositol and/or the phosphoinositide signal tranduction pathway operates as part of the communication system between chloroplasts and the nucelus ( the hypothesized chloroplast-to-nucleus signal transduction pathway in plants). The investigator will use enzyme assays, western analyses, Immunocytochemical analyses, in situ hybridizations, phospholipid studies, and molecular analyses to (1) correlate sites of inositol biosynthesis with sites of PI biosynthesis and (2) isolate mutants defective in inositol metabolism. This project is expected to yield direct evidence concerning inositol's function in chloroplast and other subcellular organelles shown to synthesize inositol. The findings will be directly applicable to the overall picture (still lacking) of the important plant phosphoinositide signal transduction pathway. This project is concerned with synthesis of inositol in plants. While much is known about inositol phosphate signaling in animals, the corresponding issues in plants have not been well studied. The overall long-term goal of the work is to understand inositol's role in the biology of subcellular organelles. The investigator has shown in previous studies that the key biosynthetic enzyme, myo-inositol-1-phosphate synthase, has temporally and spatially controlled isoforms; this unexpected finding is novel. This project will involve a detailed study of the sites of inositol and phosphionositol synthesis. A combination of immunocytochemical analyses, in situ hybridization studies, and enzyme activity measurements will be performed using Phaseolus vulgaris and Arabidopsis thaliana. This work will help in developing an understanding of cell signaling in plants, as well as of regulation of an important metabolic pathway.