Higher plants constitute one of our most important natural resources. They provide not only foodstuffs, fibers, and woods, but many chemicals, such as oils, flavorings, dyes, and pharmaceuticals. Although plants are renewable resources, some species are becoming more difficult to obtain in sufficient amounts to meet increasing demands. Destruction of natural habitats and technical difficulties in cultivation also are driving the drastic reductions in plant availability. For example, it is claimed that the demand for paclitaxel, a potent anticancer compound, could endanger forests of Taxus brevifolia (Pacific yew) because of the low paclitaxel content (40–100 mg/kg of bark) in and slow growth of the trees (1). For many natural chemicals it is possible to synthesize alternatives from petroleum, coal, or both. The economic limitations of chemical synthesis and the pollution that accompanies this type of chemical synthesis, however, have led to the development of cell culture and molecular engineering of plants for the production of important and commodity chemicals. Plant cell and organ culture offer promising alternatives for the production of chemicals because totipotency enables plant cells and organs to produce useful secondary metabolites in vitro (2). Cell culture is also advantageous in that useful metabolites are obtained under a controlled environment, independent of climatic changes and soil conditions. In addition, the products are free of microbe and insect contamination. Fermentation technology also can be used to produce desired metabolites and can be optimized to maintain high and stable yields of known quality by cellular and molecular breeding techniques to further improve productivity and quality. After extensive empirical trials, some metabolites are now being produced by large-scale cell culture (e.g., shikonin and berberine; ref. 2), but the numbers of compounds that are producible commercially by cell culture technology are still very few. The main limitations are low productivity and the necessity of the down-stream processing of the desired compounds. Molecular engineering of secondary metabolites has the potential to increase productivity and improve product composition. The Solanaceae produce a range of biologically active alkaloids that include nicotine and the tropane alkaloids (3). Tropane alkaloids, such as hyoscyamine (atropine) and scopolamine (hyoscine), which are found mainly in Hyoscyamus, Duboisia, Atropa, and Scopolia species, together with their semisynthetic derivatives, are used as parasympatholytics that competitively antagonize acetylcholine. Both the tropane ring moiety of the tropane alkaloids and the pyrrolidine ring of nicotine are derived from putrescine by way of N-methylputrescine (MP) (Fig. (Fig.1).1). Because putrescine is metabolized to polyamines such as spermidine and spermine, the N-methylation of putrescine catalyzed by putrescine N-methyltransferase (PMT) is the first committed step in the biosynthesis of these alkaloids. Figure 1 Biosynthetic pathways of tropane alkaloids and nicotine. Tropane alkaloids and nicotine are derived from diamine putrescine produced from ornithine by ornithine decarboxylase (ODC), arginine, or both (28, 29). Putrescine is N-methylated by PMT then ... Isoquinoline alkaloids are some of the major metabolites successfully produced by plant cell culture. So far, about 60 have been isolated from plant cell cultures (ref. 4 and references therein). One example is berberine, a benzylisoquinoline alkaloid obtained from Coptis (Ranunculaceae) that is used as an antibacterial agent. Berberine biosynthesis in plant cells has been well investigated at the enzyme level (5–7). The biosynthetic pathway leading from l-tyrosine to berberine has 13 different enzymatic reactions that involve a norcoclaurine synthase, an N-methyltransferase, three O-methyltransferases (OMTs), a hydroxylase, a berberine bridge enzyme, a methylenedioxy ring-forming enzyme, and a tetrahydro protoberberine oxidase (Fig. (Fig.2).2). cDNAs of several enzymes in this pathway have been isolated and characterized: norcoclaurine 6OMT (8), a hydroxylase (9), 3′-hydroxy-N-methyl-coclaurine 4′OMT (8), a berberine bridge enzyme (10), and (S)-scoulerine 9-O-methyltransferase (SMT) (11). Figure 2 Schematic biosynthetic pathway for a variety of isoquinoline alkaloids. 1, l-tyrosine decarboxylase; 2, phenolase; 3, l-tyrosine transaminase; 4, p-hydroxyphenylpyruvate decarboxylase; 5, (S)-norcoclaurine synthase; 6, (S)-adenosyl-l-methionine:norcoclaurine ... We previously reported that overexpression of hyoscyamine 6β-hydroxylase in Atropa belladonna efficiently converts this species' main alkaloid, hyoscyamine, to scopolamine (12). This successful metabolic engineering of a medicinal plant has raised prospects for biotechnological applications of secondary metabolite production, but fundamental difficulties remain in transforming the host plants (e.g., Catharanthus; ref. 13). Our recent attempts to improve the production of putrescine-derived alkaloids and isoquinoline alkaloids by molecular engineering are reported.