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BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES

Transcriptomic and Reverse Genetic Analysesof Branched-Chain Fatty Acid and Acyl Sugar Production in Solanum pennellii and Nicotiana benthamiana^1,[W],[OA]

Department of Biology, Area 7, University of York, York YO10 5YW, United Kingdom

Acyl sugars containing branched-chain fatty acids (BCFAs) are exuded by glandular trichomes of many species in Solanaceae, having an important defensive role against insects. From isotope-feeding studies, two modes of BCFA elongation have been proposed: (1) fatty acid synthase-mediated two-carbon elongation in the high acyl sugar-producing tomato species Solanum pennellii and Datura metel; and (2) -keto acid elongation-mediated one-carbon increments in several tobacco (Nicotiana) species and a Petunia species. To investigate the molecular mechanisms underlying BCFAs and acyl sugar production in trichomes, we have taken a comparative genomic approach to identify critical enzymatic steps followed by gene silencing and metabolite analysis in S. pennellii and Nicotiana benthamiana. Our study verified the existence of distinct mechanisms of acyl sugar synthesis in Solanaceae. From microarray analyses, genes associated with -keto acid elongation were found to be among the most strongly expressed in N. benthamiana trichomes only, supporting this model in tobacco species. Genes encoding components of the branched-chain keto-acid dehydrogenase complex were expressed at particularly high levels in trichomes of both species, and we show using virus-induced gene silencing that they are required for BCFA production in both cases and for acyl sugar synthesis in N. benthamiana. Functional analysis by down-regulation of specific KAS I genes and cerulenin inhibition indicated the involvement of the fatty acid synthase complex in BCFA production in S. pennellii. In summary, our study highlights both conserved and divergent mechanisms in the production of important defense compounds in Solanaceae and defines potential targets for engineering acyl sugar production in plants for improved pest tolerance.

² Present address: Molecular, Cellular, and Developmental Biology, University of Michigan, 830 North University, Ann Arbor, MI 48109.

³ Present address: U.S. Department of Agriculture-Agricultural Research Service and Boyce Thompson Institute for Plant Research, Tower Road, Cornell University, Ithaca, NY 14853.

⁴ Present address: Physiology Laboratory, Downing Street, Cambridge CB2 3EG, United Kingdom.

⁵ Present address: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom.

⁶ Present address: Samuel Roberts Noble Foundation, 2510 Sam Noble Parkway, Ardmore, OK 73401.

⁷ Present address: Nestlé R&D Center Tours, Plant Science and Technology, 101 Avenue G. Eiffel, 37390 Notre Dame d'Oé, France.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: Pierre Broun (pierre.broun{at}rdto.nestle.com).

^[W] The online version of this article contains Web-only data.

^[OA] Open Access articles can be viewed online without a subscription.

www.plantphysiol.org/cgi/doi/10.1104/pp.108.129510

^* Corresponding author; e-mail pierre.broun{at}rdto.nestle.com.

Received September 7, 2008; accepted October 13, 2008; published October 17, 2008.