Plant Molecular Biology 51: 895–911, 2003. © 2003 Kluwer Academic Publishers. Printed in the Netherlands. 895 Jasmonate biosynthesis and the allene oxide cyclase family of Arabidopsis thaliana Irene Stenzel 1 , Bettina Hause 2 , Otto Miersch 1 , Tobias Kurz 1 , Helmut Maucher 3 , Heiko Weichert 3 , Jörg Ziegler 1 , Ivo Feussner 3 and Claus Wasternack 1,∗ 1 Institute of Plant Biochemistry, Department of Natural Product Biotechnology, Weinberg 3, 06120 Halle/Saale,Germany ( ∗ author for correspondence; e-mail cwastern@ipb-halle.de); 2 Institute of Plant Biochem- istry, Department of Secondary Metabolism, Weinberg 3, 06120 Halle/Saale, Germany; 3 Institute for Plant Science and Crop Research (IPK), Department of Molecular Cell Biology, Corrensstrasse 3, 06466 Gatersleben, Germany Received 21 April 2002; accepted 19 September 2002 Key words: allene oxide cyclase family, Arabidopsis thaliana, jasmonate biosynthesis, opr3 mutant, oxylipins Abstract In biosynthesis of octadecanoids and jasmonate (JA), the naturally occurring enantiomer is established in a step catalysed by the gene cloned recently from tomato as a single-copy gene (Ziegler et al., 2000). Based on sequence homology, four full-length cDNAs were isolated from Arabidopsis thaliana ecotype Columbia coding for proteins with AOC activity. The expression of AOC genes was transiently and differentially up-regulated upon wounding both locally and systemically and was induced by JA treatment. In contrast, AOC protein appeared at constitutively high basal levels and was slightly increased by the treatments. Immunohistochemical analyses revealed abundant occurrence of AOC protein as well as of the preceding enzymes in octadecanoid biosynthesis, lipoxygenase (LOX) and allene oxide synthase (AOS), in fully developed tissues, but much less so in 7-day old leaf tissues. Metabolic profiling data of free and esterified polyunsaturated fatty acids and lipid peroxidation products including JA and octadecanoids in wild-type leaves and the jasmonate-deficient mutant OPDA reductase 3(opr3) revealed preferential activity of the AOS branch within the LOX pathway. 13-LOX products occurred predominantly as esterified derivatives, and all 13-hydroperoxy derivatives were below the detection limits. There was a constitutive high level of free 12-oxo-phytodienoic acid (OPDA) in untreated wild-type and opr3 leaves, but an undetectable expression of AOC. Upon wounding opr3 leaves exhibited only low expression of AOC, wounded wild-type leaves, however, accumulated JA and AOC mRNA. These and further data suggest regulation of JA biosynthesis by OPDA compartmentalization and a positive feedback by JA during leaf development. Abbreviations: α-LeA, α-linolenic acid; AOC, allene oxide cyclase; cet, mutant with constitutive expression of thionin; dad1, mutant with delayed anther dehiscence1; dn-OPDA, dinor-12-oxo-phytodienoic acid; 13-HPOT, 13S-hydroperoxy-(9Z,11E,15Z)-octadecatrienoic acid; JA, jasmonic acid; JAME, JA methyl ester; L.A, linoleic acid; LOX, lipoxygenase; OPDA, 12-oxo-phytodienoic acid; opr3, mutant defective in OPR3; OPR3, OPDA reductase3; PLA1, phospholipase of the A1 type; PUFA, polyunsaturated fatty acids; SA, salicylate Introduction Generation of lipid-derived signal molecules is a common phenomenon in higher organisms. In plants jasmonates and octadecanoids are of particular im- The nucleotide sequence data reported will appear in the EMBL GenBank and DDBJ Nucleotide Sequence Databases under the ac- cession numbers AJ308483 (AOC1); AJ308484 (AOC2); AJ308485 (AOC3) and AJ308486 (AOC4). portance (Bergey et al., 1996; Ryan, 2000). They originate from polyunsaturated fatty acids (PUFA) and are formed by one of the seven different branches of the LOX pathway, the AOS branch (Feussner and Wasternack, 2002). The other branches lead to leaf aldehydes and leaf alcohols as well as various deriv- atives of PUFAs such as epoxy-, hydroxy-, keto- or ether PUFA and epoxy hydroxy PUFA (Feussner and Wasternack, 2002).