title: Efficient metabolic pathway engineering in transgenic tobacco and tomato plastids with synthetic multigene operons.
abstract: The engineering of complex metabolic pathways requires the concerted expression of multiple genes. In plastids (chloroplasts) of plant cells, genes are organized in operons that are coexpressed as polycistronic transcripts and then often are processed further into monocistronic mRNAs. Here we have used the tocochromanol pathway (providing tocopherols and tocotrienols, collectively also referred to as "vitamin E") as an example to establish principles of successful multigene engineering by stable transformation of the chloroplast genome, a technology not afflicted with epigenetic variation and/or instability of transgene expression. Testing a series of single-gene constructs (encoding homogentisate phytyltransferase, tocopherol cyclase, and γ-tocopherol methyltransferase) and rationally designed synthetic operons in tobacco and tomato plants, we (i) confirmed previous results suggesting homogentisate phytyltransferase as the limiting enzymatic step in the pathway, (ii) comparatively characterized the bottlenecks in tocopherol biosynthesis in transplastomic leaves and tomato fruits, and (iii) achieved an up to tenfold increase in total tocochromanol accumulation. In addition, our results uncovered an unexpected light-dependent regulatory link between tocochromanol metabolism and the pathways of photosynthetic pigment biosynthesis. The synthetic operon design developed here will facilitate future synthetic biology applications in plastids, especially the design of artificial operons that introduce novel biochemical pathways into plants.
authors: Max-Planck-Institut für Molekulare Pflanzenphysiologie, D-14476 Potsdam-Golm,  Germany.
authors: Yinghong
authors: Y
authors: Lu
authors: Habib
authors: H
authors: Rijzaani
authors: Daniel
authors: D
authors: Karcher
authors: Stephanie
authors: S
authors: Ruf
authors: Ralph
authors: R
authors: Bock
mesh_terms: D017343:Genes, Plant; D018551:Solanum lycopersicum / Q000235:genetics*; D008969:Molecular Sequence Data; D009876:Operon; D030821:Plants, Genetically Modified / Q000235:genetics*; D018087:Plastids; D014026:Nicotiana / Q000235:genetics*; D024505:Tocopherols / Q000378:metabolism
chemical_list: D024505:Tocopherols
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