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.
