Characterization of Three Potato Lipoxygenases with Distinct Enzymatic Activities and Different Organ-specific and Wound-regulated Expression Patterns

Abstract

Lipoxygenases are ubiquitous enzymes in eukaryotes. In plants, lipoxygenases are involved in the synthesis of the hormone jasmonic acid that regulates plant responses to wounding and, in addition, is an inducer of tuberization in potato. We have isolated potato lipoxygenase cDNA clones. From their deduced amino acid sequences, three distinct classes are defined (Lox1, Lox2, and Lox3). They are encoded in gene families that display organ-specific expression, lox1 being expressed mostly in tubers and roots, lox2 in leaves, and lox3 in leaves and roots. Consistent with their organ-specific expression pattern, Lox1 expressed in bacteria preferentially uses as substrate linoleic acid, abundant in membrane lipids of tubers, whereas linolenic acid, prevalent in leaves, is the preferred substrate for the other two classes of lipoxygenase. Analyses on reaction products of the enzymes expressed in bacteria reveal that Lox1 primarily produces 9- hydroperoxides. In contrast, the jasmonic acid precursor, 13-hydroperoxylinolenic acid, is the major product of the action of Lox2 and Lox3 on linolenic acid. Upon wounding, the levels of Lox2 and Lox3 transcripts rise markedly in leaves. While Lox3 mRNA accumulation peaks as early as 30 min after wounding, Lox2 shows a steady increase over a 24-h time course, suggesting different roles for these lipoxygenase isoforms in the synthesis of the plant hormone jasmonic acid.

A Gene Encoding a Chloroplast-Targeted Lipoxygenase in Tomato Leaves Is Transiently Induced by Wounding, Systemin, and Methyl Jasmonate

“…The current picture of the LOX gene family in tomato thus resembles the situation in potato described by Royo et al (1996), with three classes identified to date on the basis of sequence similarity: the Loxl class includes TomLoxA and TomLoxB (Ferrie et al, 1994), the Lox2 class is represented by TomLoxC, and Lox3 is defined by TomLoxD. As shown for AtLOX2 (Bell et al, 1995) and suggested for rice RLL and for potato LOXZ and LOX3, the N-terminal extensions on both TomLOXC and TomLOXD might function as chloroplast transit peptides.…”

“…More recently, additional Lox genes were cloned, and TomLoxC and TomLoxD were found to be highly similar to Lox2 and Lox3, respectively (Royo et al, 1996), two potato Lox genes that appear to be the homologs of the tomato genes described here. The overall identity of TomLOXC and TomLOXD with the previously cloned TomLOXA and TomLOXB (Ferrie et al, 1994) is relatively low.…”

“…In transgenic Arabidopsis tkaliana plants having reduced levels of synthesis of the chloroplast AtLOX2, the wound-induced accumulation of JA was suppressed, and the induction of the AtVsp gene by wounding was reduced (Bell et al, 1995). As the research reported herein was being completed, three Lox cDNAs were characterized from potato tubers and leaves that were organ-specific; Loxl was expressed in tubers and roots, Lox2 was expressed in leaves, and Lox3 was expressed in leaves and roots (Royo et al, 1996). Both LOX2-and LOX3-predicted proteins exhibited a putative chloroplast leader sequence, and their mRNAs accumulated in leaves in response to wounding.…”

“…Percentage identity/similarity between cDNA-deduced amino acid sequences of plant LOXs TomLOXC and TomLOXD with TomLOXA (Ferrie et al, 1994), potato LOX2 and LOX3 (PotLOX2 and PotLOX3; Royo et al, 1996), AtLOX2 , and rice RLL (Peng et al, 1994 (Bell and Mullet, 1993), rice RLL (Peng et al, 1994), and TomLOXA (Ferrie et al, 1994). Spacings were introduced to maximize sequence identity and are represented by dots.…”

Variations in CYP74B2 (Hydroperoxide Lyase) Gene Expression Differentially Affect Hexenal Signaling in the Columbia and Landsberg erecta Ecotypes of Arabidopsis

“…Following up on our P450 microarray, we analyzed CYP71B24, CYP706A1, CYP81D10, and CYP734A1, which were loci expressed at higher levels in the Col ecotype, as well as CYP83A1, CYP706A2, CYP707A3, and chalcone synthase-like (CHS-L), which were loci expressed at higher levels in the Ler ecotype. We also analyzed expression of the PR1-similar locus, the LOX2 locus that codes for a chloroplast 13-LOX providing substrates for HPL, and the LOX1 locus that codes for a 9-LOX not generating substrates for HPL (Royo et al, 1996;Blee, 2002). As shown in Figure 6, the expression patterns from RT-PCR analyses are generally in agreement with the P450 microarray results: Nearly all genes shown by microarray analysis to be expressed higher in one ecotype were also expressed higher in that ecotype by RT-PCR analysis.…”