ABSTRACT
In plants, lipoxygenases (LOXs) play a crucial role in biotic and abiotic stresses. In our previous study, five 13-LOX genes of oriental melon were regulated by abiotic stress but it is unclear whether the 9-LOX is involved in biotic and abiotic stresses. The promoter analysis revealed that CmLOX09 (type of 9-LOX) has hormone elements, signal substances, and stress elements. We analyzed the expression of CmLOX09 and its downstream genes-CmHPL and CmAOS-in the leaves of four-leaf stage seedlings of the oriental melon cultivar "Yumeiren" under wound, hormone, and signal substances. CmLOX09, CmHPL, and CmAOS were all induced by wounding. CmLOX09 was induced by auxin (indole acetic acid, IAA) and gibberellins (GA3); however, CmHPL and CmAOS showed differential responses to IAA and GA3. CmLOX09, CmHPL, and CmAOS were all induced by hydrogen peroxide (H2O2) and methyl jasmonate (MeJA), while being inhibited by abscisic acid (ABA) and salicylic acid (SA). CmLOX09, CmHPL, and CmAOS were all induced by the powdery mildew pathogen Podosphaera xanthii. The content of 2-hexynol and 2-hexenal in leaves after MeJA treatment was significantly higher than that in the control. After infection with P. xanthii, the diseased leaves of the oriental melon were divided into four levels-levels 1, 2, 3, and 4. The content of jasmonic acid (JA) in the leaves of levels 1 and 3 was significantly higher than that in the level 0 leaves. In summary, the results suggested that CmLOX09 might play a positive role in the response to MeJA through the hydroperoxide lyase (HPL) pathway to produce C6 alcohols and aldehydes, and in the response to P. xanthii through the allene oxide synthase (AOS) pathway to form JA.
Subject(s)
Cucurbitaceae/enzymology , Hormones/metabolism , Lipoxygenase/metabolism , Plant Proteins/metabolism , Signal Transduction , Abscisic Acid , Acetates/chemistry , Aldehyde-Lyases/metabolism , Aldehydes/chemistry , Cucurbitaceae/genetics , Cyclopentanes/chemistry , Cytochrome P-450 Enzyme System/metabolism , Gene Expression Profiling , Gene Expression Regulation, Plant , Hydrogen Peroxide/metabolism , Intramolecular Oxidoreductases/metabolism , Oxylipins/chemistry , Plant Leaves/enzymology , Plant Leaves/genetics , Plant Proteins/genetics , Promoter Regions, Genetic , Salicylic Acid/chemistry , Seedlings/metabolism , Stress, Physiological , TransgenesABSTRACT
To better understand the function role of the melon CmLOX18 gene in the biosynthesis of C6 volatiles during fruit ripening, we biochemically characterized CmLOX18 and identified its subcellular localization in transgenic tomato plants. Heterologous expression in yeast cells showed that the molecular weight of the CmLOX18 protein was identical to that predicted, and that this enzyme possesseed lipoxygenase activity. Linoleic acid was demonstrated to be the preferred substrate for the purified recombinant CmLOX18 protein, which exhibited optimal catalytic activity at pH 4.5 and 30 °C. Chromatogram analysis of the reaction product indicated that the CmLOX18 protein exhibited positional specificity, as evidenced by its release of only a C-13 oxidized product. Subcellular localization analysis by transient expression in Arabidopsis protoplasts showed that CmLOX18 was localized to non-chloroplast organelles. When the CmLOX18 gene was transgenically expressed in tomato via Agrobacterium tumefaciens-mediated transformation, it was shown to enhance expression levels of the tomato hydroperoxide lyase gene LeHPL, whereas the expression levels of six TomLox genes were little changed. Furthermore, transgenic tomato fruits exhibited increases in the content of the C6 volatiles, namely hexanal, (Z)-3-hexanal, and (Z)-3-hexen-1-ol, indicating that CmLOX18 probably plays an important role in the synthesis of C6 compounds in fruits.
