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Isoprenoids are synthesized through the condensation of five-carbon intermediates, isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), derived from two distinct biosynthetic routes: cytosolic mevalonate (MVA) and plastidial 2-C-methyl-D: -erythritol 4-phosphate (MEP) pathways. 1-Hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase (IDS; EC 1.17.1.2), which catalyzes the last step of MEP pathway, was cloned as a multicopy gene from gymnosperms Ginkgo biloba (GbIDS1, GbIDS2, and GbIDS2-1) and Pinus taeda (PtIDS1 and PtIDS2), and characterized. Phylogenetic tree constructed with other plant IDSs demonstrated gymnosperm IDSs were distinctively different from angiosperm IDSs. The gymnosperm IDS clade contained two subclades, one composed of GbIDS1 and PtIDS1, and the other composed of GbIDS2, GbIDS2-1, and PtIDS2. G. biloba IDSs, except GbIDS2-1, successfully complemented Escherichia coli DLYT1, a lytB disruptant, confirming the in vivo competency of isozymes. During the 4 weeks study period, although transcript levels of GbIDS1s were similar both in roots and leaves of cultured G. biloba embryo, the transcripts of GbIDS2 predominantly occurred in the embryo roots, where diterpene ginkgolides are biosynthesized. Levels of PtIDS2 transcripts in the diterpenoid resin-producing wood were 4-5 times higher than those in other tissues. Higher levels of GbIDS1 transcripts were induced by light, whereas those of GbIDS2 were increased by methyl jasmonate treatment. These results strongly imply GbIDS2 and PtIDS2 have high correlation with secondary metabolism. In Arabidopsis transient expression system, N-terminal 100 amino acid residues of GbIDS1 delivered fused GFP protein into chloroplast as well as cytosol and nucleus, whereas those of GbIDS2, GbIDS2-1, and two PtIDSs delivered GFP only into chloroplast.

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In Hevea brasiliensis, laticifers produce and accumulate rubber particles. Despite observation using histochemical methods, development stage structure and structures with ceasing functions have rarely been described. Spectral confocal laser scanning microscopy with Nile red staining simplifies laticifer structure observation in tangential sections while enhancing the resolution. Laticifer and ray images were extracted from unmixed images and used to monitor changes during growth. A laticifer network structure developed from increased anastomoses between adjoining laticifers outside of the conducting phloem, but because of increased radial division and growth of rays, the network structure ruptured and disintegrated. We also investigated immunohistochemical localization of two rubber particle-associated proteins in the laticifers: small rubber particle protein (SRPP) and rubber elongation factor (REF). Mature bark test results show that SRPP is localized only in the laticifer layers in the conducting phloem; REF is localized in all laticifer layers. Because SRPP plays a positive role in rubber biosynthesis, results show that the rubber biosynthesis capability of laticifers is concentrated where rays and the sieve tube actively transport metabolites.