@article {2147, title = {Identification of the wax ester synthase/acyl-coenzyme A: Diacylglycerol acyltransferase WSD1 required for stem wax ester biosynthesis in Arabidopsis}, journal = {Plant Physiology}, volume = {148}, number = {1}, year = {2008}, note = {ISI Document Delivery No.: 344SMTimes Cited: 16Cited Reference Count: 52Li, Fengling Wu, Xuemin Lam, Patricia Bird, David Zheng, Huanquan Samuels, Lacey Jetter, Reinhard Kunst, Ljerka}, month = {Sep}, pages = {97-107}, type = {Article}, abstract = {Wax esters are neutral lipids composed of aliphatic alcohols and acids, with both moieties usually long-chain (C-16 and C-18) or very-long-chain (C-20 and longer) carbon structures. They have diverse biological functions in bacteria, insects, mammals, and terrestrial plants and are also important substrates for a variety of industrial applications. In plants, wax esters are mostly found in the cuticles coating the primary shoot surfaces, but they also accumulate to high concentrations in the seed oils of a few plant species, including jojoba (Simmondsia chinensis), a desert shrub that is the major commercial source of these compounds. Here, we report the identification and characterization of WSD1, a member of the bifunctional wax ester synthase/diacylglycerol acyltransferase gene family, which plays a key role in wax ester synthesis in Arabidopsis (Arabidopsis thaliana) stems, as first evidenced by severely reduced wax ester levels of in the stem wax of wsd1 mutants. In vitro assays using protein extracts from Escherichia coli expressing WSD1 showed that this enzyme has a high level of wax synthase activity and approximately 10-fold lower level of diacylglycerol acyltransferase activity. Expression of the WSD1 gene in Saccharomyces cerevisiae resulted in the accumulation of wax esters, but not triacylglycerol, indicating that WSD1 predominantly functions as a wax synthase. Analyses of WSD1 expression revealed that this gene is transcribed in flowers, top parts of stems, and leaves. Fully functional yellow fluorescent protein-tagged WSD1 protein was localized to the endoplasmic reticulum, demonstrating that biosynthesis of wax esters, the final products of the alcohol-forming pathway, occurs in this subcellular compartment.}, keywords = {ABC, CLONING, CONDENSING ENZYME, ELONGATION, GENE-EXPRESSION, PLANT CUTICULAR WAXES, SURFACES, THALIANA, TRANSFORMATION, TRANSGENIC ARABIDOPSIS, TRANSPORTER}, isbn = {0032-0889}, url = {://000258947600010}, author = {Li, F. and Wu, X. and Lam, P. and Bird, D. and Zheng, H. and Samuels, L. and Jetter, R. and Kunst, L.} } @article {2113, title = {Plant surface lipid biosynthetic pathways and their utility for metabolic engineering of waxes and hydrocarbon biofuels}, journal = {Plant Journal}, volume = {54}, number = {4}, year = {2008}, note = {ISI Document Delivery No.: 299KRTimes Cited: 12Cited Reference Count: 122Jetter, Reinhard Kunst, Ljerka}, month = {May}, pages = {670-683}, type = {Review}, abstract = {Due to their unique physical properties, waxes are high-value materials that are used in a variety of industrial applications. They are generated by chemical synthesis, extracted from fossil sources, or harvested from a small number of plant and animal species. As a result, the diversity of chemical structures in commercial waxes is low and so are their yields. These limitations can be overcome by engineering of wax biosynthetic pathways in the seeds of high-yielding oil crops to produce designer waxes for specific industrial end uses. In this review, we first summarize the current knowledge regarding the genes and enzymes generating the chemical diversity of cuticular waxes that accumulate at the surfaces of primary plant organs. We then consider the potential of cuticle biosynthetic genes for biotechnological wax production, focusing on selected examples of wax ester chain lengths and isomers. Finally, we discuss the genes/enzymes of cuticular alkane biosynthesis and their potential in future metabolic engineering of plants for the production of renewable hydrocarbon fuels.}, keywords = {ARABIDOPSIS-THALIANA, BRASSICA-OLERACEA, chain lengths, CONDENSING ENZYME, CUTICULAR WAX, cuticular waxes, ECERIFERUM MUTANTS, EPICUTICULAR WAX, ESTERS, fatty acid elongation, FATTY ACYL-COENZYME, HYDROCARBONS, industrial products, LEAVES PISUM-SATIVUM, MOLECULAR CHARACTERIZATION, SACCHAROMYCES-CEREVISIAE}, isbn = {0960-7412}, url = {://000255755000012}, author = {Jetter, R. and Kunst, L.} } @article {792, title = {The SHINE clade of AP2 domain transcription factors activates wax biosynthesis, alters cuticle properties, and confers drought tolerance when overexpressed in Arabidopsis}, journal = {Plant Cell}, volume = {16}, number = {9}, year = {2004}, note = {ISI Document Delivery No.: 854TTTimes Cited: 99Cited Reference Count: 62}, month = {Sep}, pages = {2463-2480}, type = {Article}, abstract = {The interface between plants and the environment plays a dual role as a protective barrier as well as a medium for the exchange of gases, water, and nutrients. The primary aerial plant surfaces are covered by a cuticle, acting as the essential permeability barrier toward the atmosphere. It is a heterogeneous layer composed mainly of lipids, namely cutin and intracuticular wax with epicuticular waxes deposited on the surface. We identified an Arabidopsis thaliana activation tag gain-of-function mutant shine (shn) that displayed a brilliant, shiny green leaf surface with increased cuticular wax compared with the leaves of wild-type plants. The gene responsible for the phenotype encodes one member of a clade of three proteins of undisclosed function, belonging to the plant-specific family of AP2/EREBP transcription factors. Overexpression of all three SHN clade genes conferred a phenotype similar to that of the original shn mutant. Biochemically, such plants were altered in wax composition (very long fatty acid derivatives). Total cuticular wax levels were increased sixfold in shn compared with the wild type, mainly because of a ninefold increase in alkanes that comprised approximately half of the total waxes in the mutant. Chlorophyll leaching assays and fresh weight loss experiments indicated that overexpression of the SHN genes increased cuticle permeability, probably because of changes in its ultrastructure. Likewise, SHN gene overexpression altered leaf and petal epidermal cell structure, trichome number, and branching as well as the stomatal index. Interestingly, SHN overexpressors displayed significant drought tolerance and recovery, probably related to the reduced stomatal density. Expression analysis using promoter-beta-glucuronidase fusions of the SHN genes provides evidence for the role of the SHN clade in plant protective layers, such as those formed during abscission, dehiscence, wounding, tissue strengthening, and the cuticle. We propose that these diverse functions are mediated by regulating metabolism of lipid and/or cell wall components.}, keywords = {CER MUTANTS, CHEMICAL-COMPOSITION, CONDENSING ENZYME, CUTICULAR WAX, ECERIFERUM, EPICUTICULAR WAX, EPIDERMAL-CELL DIFFERENTIATION, ORGAN FUSION, POLLEN FERTILITY, RESPONSIVE GENE-EXPRESSION, SEPARATION PROCESSES}, isbn = {1040-4651}, url = {://000223927000018}, author = {Aharoni, A. and Dixit, S. and Jetter, R. and Thoenes, E. and van Arkel, G. and Pereira, A.} } @article {1026, title = {Tomato fruit cuticular waxes and their effects on transpiration barrier properties: functional characterization of a mutant deficient in a very-long-chain fatty acid beta-ketoacyl-CoA synthase}, journal = {Journal of Experimental Botany}, volume = {55}, number = {401}, year = {2004}, note = {ISI Document Delivery No.: 829GETimes Cited: 49Cited Reference Count: 44}, month = {Jun}, pages = {1401-1410}, type = {Article}, abstract = {Cuticular waxes play a pivotal role in limiting transpirational water loss across the plant surface. The correlation between the chemical composition of the cuticular waxes and their function as a transpiration barrier is still unclear. In the present study, intact tomato fruits (Lycopersicon esculentum) are used, due to their astomatous surface, as a novel integrative approach to investigate this composition-function relationship: wax amounts and compositions of tomato were manipulated before measuring unbiased cuticular transpiration. First, successive mechanical and extractive wax-removal steps allowed the selective modification of epi- and intracuticular wax layers. The epicuticular film consisted exclusively of very-long-chain aliphatics, while the intracuticular compartment contained large quantities of pentacyclic triterpenoids as well. Second, applying reverse genetic techniques, a loss-of-function mutation with a transposon insertion in a very-long-chain fatty acid elongase beta-ketoacyl-CoA synthase was isolated and characterized. Mutant leaf and fruit waxes were deficient in n-alkanes and aldehydes with chain lengths beyond C-30, while shorter chains and branched hydrocarbons were not affected. The mutant fruit wax also showed a significant increase in intracuticular triterpenoids. Removal of the epicuticular wax layer, accounting for one-third of the total wax coverage on wild-type fruits, had only moderate effects on transpiration. By contrast, reduction of the intracuticular aliphatics in the mutant to approximately 50\% caused a 4-fold increase in permeability. Hence, the main portion of the transpiration barrier is located in the intracuticular wax layer, largely determined by the aliphatic constituents, but modified by the presence of triterpenoids, whereas epicuticular aliphatics play a minor role.}, keywords = {ARABIDOPSIS-THALIANA, BARRIER, BICOLOR L MOENCH, CITRUS LEAF CUTICLES, CONDENSING ENZYME, CONDUCTANCE, cuticle, ECERIFERUM CER MUTANTS, EPICUTICULAR WAX, EPICUTICULAR WAXES, EPIDERMAL, intracuticular wax, PRUNUS-LAUROCERASUS L, tomato, transpiration, TRANSPORT-PROPERTIES, WATER PERMEABILITY}, isbn = {0022-0957}, url = {://000222034600015}, author = {Vogg, G. and Fischer, S. and Leide, J. and Emmanuel, E. and Jetter, R. and Levy, A. A. and Riederer, M.} }