9514177 Cramer Plants produce a vast array of isoprenoid compounds (>10,000) that function in growth and development, electron transport, photosynthesis, anti-oxidant protection, reproduction, and disease resistance. The appropriate spatial and temporal production of specific isoprenoid endproducts from common central-pathway intermediates poses a significant regulatory challenge. The proposed reserch will address whether the plant isoprenoid pathway is organized in distinct metabolic units or "metabolons" as a mechanism to direct pathway flux into specific endproducts. In Solanaceous plants, the HMG1 isoform of 3-hydroxy-3methylglutaryl CoA reductase (HMGR, the rate-limiting enzyme producing mevalonic acid) and squalene synthetase (the key branch enzyme committing farnesyl intermediates to sterol synthesis) are coordinately regulated in association with sterol biosynthesis. Data from a variety of experimental approaches suggest that, in addition to differential regulation at the gene level, protein-protein interaction and/or subcellular localization of specific HMGRs may be important in partitioning intermediates into production of specific isoprenoids. In this project, the hypothesis that tomato HMG1 and HMG2 function in physiologically distinct pathways or metabolons leading to production of phytosterols or phytoalexins, respectively, will be tested. HMGR isozyme levels will be altered by introducing hmg1 or hmg2 sequences regulated by either constitutive or defense-inducible promoters. The HMG1 and HMG2 transgene projects will be differentially tagged to facilitate immunolocalization studies and assessment of protein stability. Many plantspecific isoprenoids have significant value for agricultural, industrial, and pharmaceutical applications (e.g., natural pesticides, rubber, and taxol). The proposed experiments will provide significant new information critical for understanding the complex biochemical and cellular mechanisms involved in isoprenoid biosynthesis.
