Jasmonic Acid Levels Are Reduced in COMATOSE ATP-Binding Cassette Transporter Mutants. Implications for Transport of Jasmonate Precursors into Peroxisomes

Abstract

We provide evidence that the peroxisomal ATP-binding cassette (ABC) transporter COMATOSE (CTS) is involved in the biosynthesis of jasmonic acid (JA) in Arabidopsis (Arabidopsis thaliana) leaves. Basal JA levels were greatly reduced but not completely abolished in two cts mutant alleles, and JA production in response to wounding showed slower kinetics and reached a lower level than wild type. We propose the operation of two parallel pathways for peroxisomal import of jasmonate precursors, a route that requires CTS function and a parallel leak pathway involving anion trapping. It is well established that the bioactive, lipid-derived compounds collectively known as jasmonates play important roles in plant defense, metabolism, and development (Farmer and Ryan, 1992; Creelman and Mulpuri, 2002). A biosynthetic pathway for JA was proposed in 1983 by Vick and Zimmerman (1983), in which JA is formed from the unsaturated fatty acid, linolenic acid (18:3), an abundant octadecanoid in higher plant membranes. In Arabidopsis, a parallel, hexadecanoid pathway based on the 16:3 fatty acid is also present (Weber et al., 1997). Most of the enzymes involved in the 18:3 pathway have now been identified by a combination of classic biochemical and genetic approaches (Creelman and Mulpuri, 2002; Turner et al., 2002). Following phospholipase-dependent release of 18:3 from chloroplast membrane lipids, molecular oxygen is introduced by 13-lipoxygenase activity to generate 13-hydroperoxy octadecatrienoic acid. Allene oxide synthase catalyzes the formation of an unstable intermediate, 12,13-epoxy-octadecatrienoic acid, which is converted to (9S,13S)-12-oxo-phytodienoic acid (OPDA) by allene oxide cyclase. The synthesis of OPDA occurs in the chloroplast, and in Arabidopsis more than 90% of OPDA is esterified to chloroplast galactolipids (Stelmach et al., 2001). In subsequent steps, OPDA is reduced to 3-oxo-2-(2′-pentenyl)-cyclopentane-1-octanoic acid (OPC:8), which is then shortened by three rounds of peroxisomal β-oxidation to yield JA. The involvement of specific isoforms of acyl-CoA oxidase (ACX1) and thiolase (KAT2) in the β-oxidation steps of wound-induced leaf JA biosynthesis has recently been demonstrated (Cruz Castillo et al., 2004). Thus, JA biosynthesis is initiated in the chloroplast and terminates in the peroxisome. Although Arabidopsis has at least three isoforms of 12-oxo-phytodienoate reductase, only OPR3 reduces the 9S,13S-stereoisomer of OPDA, the biological precursor of JA (Schaller et al., 2000). Since OPR3 is located in the peroxisome (Strassner et al., 2002), it is clear that OPDA or other intermediates in JA synthesis must be transported from the chloroplast to the peroxisome, but the nature of these transport steps is unknown. It has generally been assumed that OPDA is freely membrane permeable because of its structural similarity to fatty acids, but since OPDA is an important signaling molecule in its own right (Blechert et al., 1999; Stintzi et al., 2001), it is plausible that control of its movement between subcellular compartments would be highly regulated (Stenzel et al., 2003). This could implicate membrane-bound transporters in the chloroplast and/or peroxisome.