Duplicate maize 13-lipoxygenase genes are differentially regulated by circadian rhythm, cold stress, wounding, pathogen infection, and hormonal treatm

Most plant oxylipins, a large class of diverse oxygenated polyunsaturated fatty acids and their derivatives, are produced through the lipoxygenase (LOX) pathway. Recent progress in dicots has highlighted the biological roles of oxylipins in plant defence responses to pathogens and pests. By contrast, the physiological function of LOXs and their metabolites in monocots is poorly understood. Two maize LOXs, ZmLOX10 and ZmLOX11 that share >90% amino acid sequence identity but are localized on different chromosomes, were cloned and characterized. Phylogenetic analysis revealed that ZmLOX10 and ZmLOX11 cluster together with well-characterized plastidic type 2 linoleate 13-LOXs from diverse plant species. Regio-specificity analysis of recombinant ZmLOX10 protein overexpressed in Escherichia coli proved it to be a linoleate 13-LOX with a pH optimum at pH 8.0. Both predicted proteins contain putative transit peptides for chloroplast import. ZmLOX10 was preferentially expressed in leaves and was induced in response to wounding, cold stress, defence-related hormones jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA), and inoculation with an avirulent strain of Cochliobolus carbonum. These data suggested a role for this gene in maize adaptation to abiotic stresses and defence responses against pathogens and pests. ZmLOX11 was preferentially expressed in silks and was induced in leaves only by ABA, indicating its possible involvement in responses to osmotic stress. In leaves, mRNA accumulation of ZmLOX10 is strictly regulated by a circadian rhythm, with maximal expression coinciding temporally with the highest photosynthetic activity. This study reveals the evolutionary divergence of physiological roles for relatively recently duplicated genes. Possible physiological functions of these 13-LOXs are suggested.

Source: J Exp Bot. (2006) vol. 57, 3767-3779

Expression of yeast SOD2 in transgenic rice results in increased salt tolerance

Agricultural productivity is severely affected by soil salinity. One possible mechanism by which plants could survive salt stress is to remove sodium ions from the cytosol via plasma membrane Na+/H+ antiporters. In the present study, we demonstrated that expressing the plasma membrane Na+/H+ antiporter SOD2 from yeast (Schizosaccharomyces pombe) in transgenic rice increased salt tolerance. These transgenic plants accumulated more K+, Ca2+, Mg2+ and less Na+ in their shoots compared with those of non-transformed controls. Moreover, measurements on isolated plasma membrane vesicles derived from the SOD2 transgenic rice plant roots showed that the vesicles had enhanced P-ATPase hydrolytic activity. Furthermore, the transformed rice plants maintained higher levels of photosynthesis and root proton exportation capacity, whereas reduced ROS generation. Physiological analysis suggested that transgenic rice plants might employ multiple mechanisms to improve their salt tolerance under salt stress conditions.

Source: Plant science (2006) vol. 170, p. 216-224

Identification of a peroxisomal acyl-activating enzyme involved in the biosynthesis of jasmonic acid in Arabidopsis

Jasmonic acid (JA) is a lipid-derived signal that regulates a wide variety of developmental and defense-related processes in higher plants. JA is synthesized from linolenic acid via an enzymatic pathway that initiates in the plastid and terminates in peroxisomes. The C18 JA precursor 12-oxo-phytodienoic acid (OPDA) is converted in the peroxisome to 3-oxo-2-(2'-[Z]-pentenyl)cyclopentane-1-octanoic acid (OPC-8:0), which subsequently undergoes three rounds of beta-oxidation to yield JA. Although most JA biosynthetic enzymes have been identified, several key steps in the pathway remain to be elucidated. To address this knowledge gap, we employed co-expression analysis to identify genes that are coordinately regulated with known JA biosynthetic components in Arabidopsis. Among the candidate genes uncovered by this approach was a 4-coumarate-CoA ligase-like member of the acyl-activating enzyme (AAE) gene family, which we have named OPC-8:0 CoA Ligase1 (OPCL1). In response to wounding, opcl1 null mutants exhibited reduced levels of JA and hyperaccumulation of OPC-8:0. Recombinant OPCL1 was active against both OPDA and OPC-8:0, as well as medium-to-long straight-chain fatty acids. Subcellular localization studies with green fluorescent protein-tagged OPCL1 showed that the protein is targeted to peroxisomes. These findings establish a physiological role for OPCL1 in the activation of JA biosynthetic precursors in leaf peroxisomes, and further indicate that OPC-8:0 is a physiological substrate for the activation step. The results also demonstrate the utility of co-expression analysis for identification of factors that contribute to jasmonate homeostasis.

Source: J Biol Chem. (2006) vol. 281, p. 33511-33520

Lipoxygenases during Brassica napus seed germination

The peroxidation of polyunsaturated fatty acids is mostly catalyzed by members of the lipoxygenase enzyme family. Lipoxygenase products can be metabolized further in the oxylipin pathway and are known as signalling substances that play a role in plant development as well as in plant responses to wounding and pathogen attack. Apart from accumulating data in model plants like Arabidopsis, information on the relevance of lipid peroxide metabolism in the crop plant oilseed rape is scarce. Thus we aimed to analyze lipoxygenases and oxylipin patterns in seedlings of oilseed rape. RNA isolated from 3 day etiolated seedlings contains mRNAs for at least two different lipoxygenases. These have been cloned as cDNAs and named Bn-Lox-lfl and Bn-Lox-2fl. The protein encoded by Bn-Lox-2fl was identified as a 13-lipoxygenase by expression in Escherichia coli. The Bn-Lox-lfl yielded an inactive protein when expressed in E. coli. Based on Bn-Lox-lfl active site determinants and on sequence homology the Bn-Lox-lfl is most likely a 9-lipoxygenase. Both genes are expressed in light-grown and etiolated cotyledons as well as in leaves. Bn-Lox-2fl protein is more abundant in cotyledons of etiolated seedlings than in cotyledons of green seedlings. Both 13- and 9-lipoxygenase-derived hydroperoxides can be detected during germination. Etiolated seedlings contain more lipoxygenase-derived hydroperoxides in non esterified fatty acids than green seedlings. The 13-lipoxygenase derivatives are 6-8-fold more abundant than the 9-derivatives. Lipoxygenase-derived hydroperoxides in esterified lipids are almost not present during germination. These results suggest that 13-lipoxygenases acting on free fatty acids dominate during B. napus seed germination.

Source: Phytochemistry (2006) vol. 67, p. 2030-2040

Over-expression of chloroplastic lipoxygenase RCI1 causes PR1 transcript accumulation in transiently transformed rice

The acquired resistance state of rice plants, rendering them resistant to subsequent infections with virulent pathogens, can be triggered by treatments with non-host pathogens, necrotizing pathogens and certain chemicals. The 13-lipoxygenase RCI1 (LOX2:Os:2) is induced specifically following treatment with chemical inducers of resistance. Here, we report that the over-expression of RCI1 in a transient transformation assay leads to PR1 transcript accumulation in rice leaves. In addition, we show that this property is due to the enzymatic activity of the transgenic protein. Furthermore, exogenous application of (13S,92,11E,15Z)-13-hydroxy-9,11,15-octadecatrienoic acid (13-HOT), an oxylipin deriving from the reductase branch of the lipoxygenase pathway, is capable of inducing PR1. These results support a role for RCI1 in the generation of signalling molecules during the establishment of the acquired resistance response in rice.

Source: Physiological and molecular plant pathology (2004) vol. 64, p. 37-43

The outcomes of concentration-specific interactions between salicylate and jasmonate signaling include synergy, antagonism, and oxidative...

The outcomes of concentration-specific interactions between salicylate and jasmonate signaling include synergy, antagonism, and oxidative stress leading to cell death

Salicylic acid (SA) has been proposed to antagonize jasmonic acid (JA) biosynthesis and signaling. We report, however, that in salicylate hydroxylase-expressing tobacco (Nicotiana tabacum) plants, where SA levels were reduced, JA levels were not elevated during a hypersensitive response elicited by Pseudomonas syringae pv phaseolicola. The effects of cotreatment with various concentrations of SA and JA were assessed in tobacco and Arabidopsis (Arabidopsis thaliana). These suggested that there was a transient synergistic enhancement in the expression of genes associated with either JA (PDF1.2 [defensin] and Thi1.2 [thionin]) or SA (PR1 [PR1a--glucuronidase in tobacco]) signaling when both signals were applied at low (typically 10–100 µM) concentrations. Antagonism was observed at more prolonged treatment times or at higher concentrations. Similar results were also observed when adding the JA precursor, -linolenic acid with SA. Synergic effects on gene expression and plant stress were NPR1- and COI1-dependent, SA- and JA-signaling components, respectively. Electrolyte leakage and Evans blue staining indicated that application of higher concentrations of SA + JA induced plant stress or death and elicited the generation of apoplastic reactive oxygen species. This was indicated by enhancement of hydrogen peroxide-responsive AoPR10--glucuronidase expression, suppression of plant stress/death using catalase, and direct hydrogen peroxide measurements. Our data suggests that the outcomes of JA-SA interactions could be tailored to pathogen/pest attack by the relative concentration of each hormone.

Source: Plant Physiology (2006) vol. 140, p. 249-262

Enhanced drought tolerance of transgenic rice plants expressing a pea manganese superoxide dismutase

We investigated the role that manganese superoxide dismutase (MnSOD), an important antioxidant enzyme, may play in the drought tolerance of rice. MnSOD from pea (Pisum sativum) under the control of an oxidative stress-inducible SWPA2 promoter was introduced into chloroplasts of rice (Oryza sativa) by Agrobacterium-mediated transformation to develop drought-tolerant rice plants. Functional expression of the pea MnSOD in transgenic rice plants (T1) was revealed under drought stress induced by polyethylene glycol (PEG) 6000. After PEG treatment the transgenic leaf slices showed reduced electrolyte leakage compared to wild type (WT) leaf slices, whether they were exposed to methyl viologen (MV) or not, suggesting that transgenic plants were more resistant to MV- or PEG-induced oxidative stress. Transgenic plants also exhibited less injury, measured by net photosynthetic rate, when treated with PEG. Our data suggest that SOD is a critical component of the ROS scavenging system in plant chloroplasts and that the expression of MnSOD can improve drought tolerance in rice.

Source: J Plant Physiol. (2005) vol. 162, p. 465-472

OsCDPK13, a calcium-dependent protein kinase gene from rice, is induced by cold and gibberellin in rice leaf sheath

Calcium-dependent protein kinases (CDPKs) play an important role in rice signal transduction, but the precise role of each individual CDPK is still largely unknown. Recently, a full-length cDNA encoding OsCDPK13 from rice seedling was isolated. To characterize the function of OsCDPK13, its responses to various stresses and hormones were analyzed in this study. OsCDPK13 accumulated in 2-week-old leaf sheath and callus, and became phosphorylated in response to cold and gibberellin (GA). OsCDPK13 gene expression and protein accumulation were up-regulated in response to GA3 treatment, but suppressed in response to abscisic acid and brassinolide. Antisense OsCDPK13 transgenic rice lines were shorter than the vector control lines, and the expression of OsCDPK13 was lower in dwarf mutants of rice than in wild type. Furthermore, OsCDPK13 gene expression and protein accumulation were enhanced in response to cold, but suppressed under salt and drought stresses. Sense OsCDPK13 transgenic rice lines had higher recovery rates after cold stress than vector control rice. The expression of OsCDPK13 was stronger in cold-tolerant rice varieties than in cold-sensitive ones. The results suggest that OsCDPK13 might be an important signaling component in the response of rice to GA and cold stress.

Source: Plant Mol Biol. (2004) vol. 55, p. 541-552