The Arabidopsis extensin gene is developmentally regulated, is induced by wounding, methyl jasmonate, abscisic and salicylic acid, and codes for a protein with unusual motifs

A single-copy extensin gene (atExt1) has been isolated from Arabidopsis thaliana (L.) Heynh. The deduced amino acid sequence consists of 374 amino acids which are organised into highly ordered repeating blocks in which Ser(Pro)₄ and Ser(Pro)₃ motifs alternate. Two copies of the Tyr-X-Tyr-Lys motif and 13 copies of the Val-Tyr-Lys motif are present, showing that this extensin may be highly cross-linked, possessing the capacity for both intra and inter-molecular bond formation. The gene atExt1 is normally expressed in the root and is silent in the leaf; wounding reverses this pattern, turning on the gene in the leaf and repressing it in the root. The promoter contains motifs which have been found to activate plant defence genes in response to salicylic acid, abscisic acid and methyl jasmonate; when these compounds are applied to the roots, the atExt1 gene is activated in the leaf. Received: 11 September 1998 / Accepted: 20 December 1998     

A novel pepper membrane-located receptor-like protein gene CaMRP1 is required for disease susceptibility, methyl jasmonate insensitivity and salt tolerance

Plant receptor proteins are involved in the signaling networks required for defense against pathogens. The novel pepper pathogen-induced gene CaMRP1 was isolated from pepper leaves infected with Xanthomonas campestris pv. vesicatoria (Xcv). This gene is predicted to encode a membrane-located receptor-like protein that has an N-terminal signal peptide and a C-terminal transmembrane helix. A CaMRP1-GFP fusion protein localized primarily to the plasma membrane of plant cells. Strong and early induction of CaMRP1 expression occurred following exposure of pepper plants to Xcv, Colletotricum coccodes, methyl jasmonate (MeJA) and wounding stress. Virus-induced gene silencing (VIGS) of CaMRP1 in pepper conferred enhanced basal resistance to Xcv infection, accompanied by induction of genes encoding basic PR1 (CaBPR1), defensin (CaDEF1) and SAR8.2 (CaSAR82A). In contrast, CaMRP1 overexpression (OX) in transgenic Arabidopsis plants resulted in increased disease susceptibility to Hyaloperonospora parasitica infection. Arabidopsis plants overexpressing CaMRP1 exhibited insensitivity to MeJA by causing reduced expression of MeJA-responsive genes. Overexpression also resulted in tolerance to NaCl and during salt stress, the expression of several abscisic acid-responsive genes was induced. Together, these results suggest that pepper CaMRP1 may belong to a new subfamily of membrane-located receptor-like proteins that regulate disease susceptibility, MeJA-insensitivity and salt tolerance.     

Accumulation of chloroplast-targeted lipoxygenase in passion fruit leaves in response to methyl jasmonate

Wounding caused local and systemic induction of lipoxygenase (LOX) activity in passion fruit (Passiflora edulis f. flavicarpa) leaves, while exposing intact plants to methyl jasmonate (MJ) vapor provoked a much stronger response. Western blot analysis of these leaf protein extracts using polyclonal antibodies against cucumber LOX, revealed an accumulation of a 90 kDa protein, consistent with LOX enzymatic assays. The inducible LOX was purified to apparent homogeneity, and in vitro analysis of LOXactivity using linoleic acid as substrate showed that it possesses C-13 specificity. Immunocytochemical localization studies using leaf tissue from MJ-treated plants demonstrated that the inducible LOX was compartmented in large quantities in the chloroplasts of mesophyll cells, associated with the stroma. The results suggest that the wound response in passion fruit plants may be mediated by a chloroplast 13-LOX, a key enzyme of the octadecanoid defense-signaling pathway.     

Interaction of methyl jasmonate, wounding and fungal elicitation during sesquiterpene induction in Hyoscyamus muticus in root cultures

The ability of methyl jasmonate (MeJa) to induce sesquiterpene production in root cultures of Hyoscyamus muticus has been studied. Although MeJa alone could not induce sesquiterpene in unwounded culture, MeJa added in the presence of wounding displayed a dose-dependent response, saturating at 50 μM. The ability to respond to MeJa declined with an increase in time between MeJa contact and wounding; however, responsiveness could be recovered by re-wounding of tissue prior to MeJa contact, suggesting that additional signaling related to wounding is required for sesquiterpene pathway induction. The saturation level of sesquiterpene induction with fungal elicitor was four times higher than the saturation level achieved by MeJa, with clear differences in sesquiterpene composition. Fungal elicitation results in a higher level of lubimin and a lower level of solavetivone production; whereas, methyl jasmonate induces predominantly solavetivone and little or no lubimin production. This suggests that fungal elicitation induces enzymes further down the sesquiterpene pathway which are not affected by MeJa. The induction of roots in contact with subsaturated levels of elicitor can be enhanced to saturation production levels by the addition of small amounts of MeJa (5–10 μmoles/l). In these studies, MeJa was consistently found to favor the earlier metabolite (solavetivone), while fungal elicitation promoted conversion to subsequent metabolites in the pathway (lubimin). The interactive role of MeJa in signal transduction for secondary metabolic production is discussed. Received: 8 June 1997 / Revision received: 13 August 1997 / Accepted: 13 September 1997     

Regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase by wounding and methyl jasmonate

HMGR (3-hydroxy-3-methylglutaryl-coenzyme A reductase; E.C.1.1.1.34) supplies mevalonate for the synthesis of many plant primary and secondary metabolites, including the terpenoid component of indole alkaloids. Suspension cultures of Camptotheca acuminata and Catharanthus roseus, two species valued for their anticancer indole alkaloids, were treated with the elicitation signal transducer methyl jasmonate (MeJA). RNA gel blot analysis from MeJA treated cultures showed a transient suppression of HMGR mRNA, followed by an induction in HMGR message. Leaf disks from transgenic tobacco plants containing a chimeric hmgl::GUS construct were also treated with MeJA and showed a dose dependent suppression of wound-inducible GUS activity. The suppression of the wound response by MeJA was limited to the first 4 h post-wounding, after which time MeJA application had no effect. The results are discussed in relation to the differential regulation of HMGR isogenes in higher plants.Abbreviations GUS -glucuronidase - hmg gene of hmgr - HMGR 3-hydroxy-3-methylglutaryl-coenzyme A reductase - JA jasmonic acid - MeJA methyl jasmonate - MUG methylumbelliferyl--d-glucuronide - TDC tryptophan decarboxylase - SDS sodium dodecyl sulfate - SS strictosidine synthase     

Dual positional specificity and expression of non-traditional lipoxygenase induced by wounding and methyl jasmonate in maize seedlings

Lipoxygenases (LOXs) catalyze the formation of fatty acid hydroperoxides involved in responses to stresses. This study examines the expression of a non-traditional dual positional specific maize LOX in response to wounding or methyl jasmonate (MeJA). Full-length maize LOX cDNA was expressed in Escherichia coli, and recombinant LOX was purified and characterized enzymatically. RP-HPLC and GC-MS analysis showed that the purified LOX converts alpha-linolenic acid into 13-hydroperoxylinolenic acid and 9-hydroperoxylinolenic acid in a 6:4 ratio. LOX mRNA accumulated rapidly and transiently in response to wounding reaching a peak of expression about 3 h after wounding. This increase followed an initial increase in endogenous jasmonic acid (JA) 1 h after wounding (JA burst). However, the expression of LOX induced by MeJA lasted longer than the expression induced by wounding, and the MeJA-induced expression seemed to be biphasic pattern composed of early and late phases. The expression of LOX in the presence of inhibitors of JA biosynthesis was not completely inhibited, but delayed in wound response and the expression period was shortened in MeJA response. These results suggest that wound-responsive JA burst may trigger the early phase of LOX expression which facilitates biosynthesis of endogenous JA through its 13-LOX activity, and subsequently leads to the activation of the late phase LOX expression in MeJA-treated maize seedlings. Implications of dual positional specificity of maize LOX in the observed expression kinetics are discussed.     

Metabolic profiling and enhanced production of phytosterols by elicitation with methyl jasmonate and silver nitrate in whole plant cultures of Lemna paucicostata

In this study, the effects of methyl jasmonate (MJ) and silver nitrate (SN) treatment on metabolic profiles and yields of phytosterols such as campesterol, stigmasterol, and β-sitosterol in whole plant cultures of Lemna paucicostata were investigated using gas chromatography–mass spectrometry coupled with multivariate statistical analysis. The MJ and SN treatments retarded the growth of L. paucicostata plants, while they enhanced the yields of three phytosterols, compared to control. Higher yields of phytosterols were attained at day 28 compared to day 42. Moreover, stigmasterol yield was the highest at 0.85 mg/g from day 28 plants grown under MJ + SN co-treated culture. Among the various metabolites, the levels of palmitic and stearic acids, which might participate in a defense mechanism, were higher in the MJ + SN condition than in control. To determine the optimal timing of MJ + SN addition, MJ + SN was added on days 21, 28, and 35 after inoculation. The total yield and productivity of phytosterol reached maximum levels when the MJ + SN was added at day 35. The highest productivity of stigmasterol (6.08 mg/L) was also achieved when MJ + SN was added on day 35.     

The level of flavonoids and amines in de-etiolated and methyl jasmonate treated seedling of common buckwheat

The effects of atmospheric methyl jasmonate on the level of flavonoids and biogenic amines in de-etiolated seedlings of common buckwheat (Fagopyrum esculentum Moench) were investigated. In cotyledons and hypocotyls of etiolated seedlings, some traces of anthocyanins were found, with no flavones and flavonols identified. A measurable content of flavones and flavonols was, however, determined in roots. De-etiolation process stimulated the accumulation of all flavonoid types. Methyl jasmonate clearly decreased the content of anthocyanins in the hypocotyl, not affecting their level in cotyledons. In case of roots, the content of anthocyanins increased after a 4-day treatment. In general, reduction in the level of flavones and flavonols was recorded only in the hypocotyl, however it was not always significant. Cotyledons of the seedlings treated with methyl jasmonate responded by a slight increase in flavonoids level. Methyl jasmonate considerably induced the accumulation of 2-phenylethylamine in all the seedling organs, increasing the content of putrescine and tryptamine in cotyledons, and decreasing the level of tryptamine in roots.     

A gene encoding a chloroplast-targeted lipoxygenase in tomato leaves is transiently induced by wounding, systemin, and methyl jasmonate.

We investigated the relationship between the expression of lipoxygenase (LOX) genes and the systemin-dependent wound response in tomato (Lycopersicon esculentum) leaves. A polymerase chain reaction-based approach was used to isolate two tomato Lox cDNAs, called TomLoxC and TomLoxD. Both TomLOXC and TomLOXD amino acid sequences possess an N-terminal extension of about 60 residues that were shown by in vitro uptake to function as transit peptides, targeting these proteins into the chloroplast. Within 30 to 50 min following wounding or systemin or methyl jasmonate treatments, the TomLoxD mRNA level increased and reached a maximum between 1 and 2 h. TomLoxC mRNA was not detectable in leaves and was not found following wounding, but it was found in ripening fruits, indicating that the two tomato Lox genes are regulated in different tissues by different processes. The results suggest that the TomLoxD gene is up-regulated in leaves in response to wounding and encodes a chloroplast LOX that may play a role as a component of the octadecanoid defense-signaling pathway.     

Inhibitory effect of methyl jasmonate and its related compounds on kinetin-induced retardation of oat leaf senescence

The senescence-promoting activities of methyl jasmonate and its related compounds were compared with respect to structure-activity relationships. The activities were assayed by using oat ( A vena saliva L. cv. Victor) leaf segments in the presence of 2 μg/ml kinetin. Dextrorotatory methyl jasmonate prepared from an authentic sample of the racemate mixture was less active than the naturally occurring levorotatory form especially at its low concentrations (0.1 to 2.5 μg/ml). The activity of jasmonic acid, the free acid form of methyl jasmonate, was much less than the methyl ester, and this relationship was true for the other compounds tested. The reduction of the unsaturated bond in the substituent at the C-2 position and the keto group at the C-3 position greatly reduced the activity. The length of the n -alkyl substituents at the C-2 position had also a significant effect on the activity. From these results, it is concluded that the important functional groups for the high senescence-promoting activity of the methyl jasmonate related compounds are the methyl acetate substituent at the C-l position, the 2' cis -pentenyl or n -pentyl group at the C-2, position and the keto group at the C-3 position in methyl jasmonate.     

Proteomics of Arabidopsis redox proteins in response to methyl jasmonate

Protein redox regulation is increasingly recognized as an important switch of protein activity in yeast, bacteria, mammals and plants. In this study, we identified proteins with potential thiol switches involved in jasmonate signaling, which is essential for plant defense. Methyl jasmonate (MeJA) treatment led to enhanced production of hydrogen peroxide in Arabidopsis leaves and roots, indicating in vivo oxidative stress. With monobromobimane (mBBr) labeling to capture oxidized sulfhydryl groups and 2D gel separation, a total of 35 protein spots that displayed significant redox and/or total protein expression changes were isolated. Using LC–MS/MS, the proteins in 33 spots were identified in both control and MeJA-treated samples. By comparative analysis of mBBr and SyproRuby gel images, we were able to determine many proteins that were redox responsive and proteins that displayed abundance changes in response to MeJA. Interestingly, stress and defense proteins constitute a large group that responded to MeJA. In addition, many cysteine residues involved in the disulfide dynamics were mapped based on tandem MS data. Identification of redox proteins and their cysteine residues involved in the redox regulation allows for a deeper understanding of the jasmonate signaling networks.     

Influence of methyl jasmonate on podophyllotoxin and 6-methoxypodophyllotoxin accumulation in Linum album cell suspension cultures

The accumulation of podophyllotoxin (PTOX) and 6-methoxypodophyllotoxin (6MPTOX) was enhanced about twofold in the suspension culture of Linum album line 2-5 aH following the addition of methyl jasmonate (MeJas) to the cultivation medium, reaching 7.69±1.45 mg/g dry weight and 1.11±0.09 mg/g dry weight, respectively. There was no increase in 6MPTOX accumulation following the addition of MeJas to suspension cells of L. album line X4SF, whereas PTOX accumulation was enhanced about tenfold to 0.49±0.10 mg/g dry weight. Phenylalanine ammonia-lyase activity increased immediately after the addition of MeJas to a cell suspension culture of line X4SF, reaching a maximum between 4 h and 1 day after elicitation, while cinnamyl alcohol dehydrogenase activity and the lignin content of the cells were not affected.     

Calcium changes and the response to methyl jasmonate in rice lodicules during anthesis

Summary. Potassium pyroantimonate precipitation was used to locate loosely bound calcium in rice (Oryza sativa L.) lodicules before and after anthesis, and flowering of panicles was accelerated by treatment with methyl jasmonate. From 1 day to 4 h before anthesis, the number of calcium precipitates in the cell walls and vacuole membranes decreased gradually, whereas they increased remarkably in the cytoplasm and nucleolus. At the beginning of anthesis, the number of calcium granules in lodicules reduced sharply, but there was a large accumulation of flocculent precipitates in the vacuoles. After anthesis, the flocculent precipitates decreased in number until they disappeared, whereas the granular precipitates started to accumulate once again. The rice florets treated with 2 mM methyl jasmonate were induced to open within 10–30 min and they then closed 0.5–1 h later. The nucleolus, cytoplasm, and vacuole membrane of the lodicule cells contained many calcium granules during flowering, although the cell walls lacked calcium. At 1 h after treatment, the number of calcium granules had decreased, while flocculent precipitates were regularly observed in the nondegenerated cells. At 6 h after treatment, calcium grains started to reappear in the cell walls. These changes in calcium precipitates before and after anthesis indicate that the opening and closing of florets correlates with the calcium level in lodicule cells. In addition, excised panicles, with florets judged to be nearing anthesis, were soaked in 2–200 mM EGTA solution for 2 min after treatment with 2 mM methyl jasmonate. The results indicate that EGTA had an antagonistic effect on the methyl jasmonate-induced floret opening in rice. Correspondence and reprints: Key Laboratory of the Ministry of Education for Plant Developmental Biology, College of Life Sciences, Wuhan University, Wuhan 430072, People’s Republic of China.     

Chemical compositions of exudates from succulent plants induced by methyl jasmonate

Methyl jasmonate (JA-Me) promoted leaf abscission of succulent plant, Crassula lycopodioides, and exudation from stems and leaves after leaf abscission when it was sprayed in whole plants. Major component of the exudate was identified as sucrose based on the results described below.

Proteomics of methyl jasmonate induced defense response in maize leaves against Asian corn borer

Background

Jasmonic acid (JA) and methyl jasmonate (MeJA) regulate plant development, resistance to stress, and insect attack by inducing specific gene expression. However, little is known about the mechanism of plant defense against herbivore attack at a protein level. Using a high-resolution 2-D gel, we identified 62 MeJA-responsive proteins and measured protein expression level changes.

Results

Among these 62 proteins, 43 proteins levels were increased while 11 proteins were decreased. We also found eight proteins uniquely expressed in response to MeJA treatment. Data are available via ProteomeXchange with identifier PXD001793. The proteins identified in this study have important biological functions including photosynthesis and energy related proteins (38.4%), protein folding, degradation and regulated proteins (15.0%), stress and defense regulated proteins (11.7%), and redox-responsive proteins (8.3%). The expression levels of four important genes were determined by qRT-PCR analysis. The expression levels of these proteins did not correlate well with their translation levels. To test the defense functions of the differentially expressed proteins, expression vectors of four protein coding genes were constructed to express in-fusion proteins in E. coli. The expressed proteins were used to feed Ostrinia furnacalis, the Asian corn borer (ACB). Our results demonstrated that the recombinant proteins of pathogenesis-related protein 1 (PR1) and thioredoxin M-type, chloroplastic precursor (TRXM) showed the significant inhibition on the development of larvae and pupae.

Conclusions

We found MeJA could not only induce plant defense mechanisms to insects, it also enhanced toxic protein production that potentially can be used for bio-control of ACB.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1363-1) contains supplementary material, which is available to authorized users.     

Fungal elicitor preparations and methyl jasmonate enhance rosmarinic acid accumulation in suspension cultures of Coleus blumei

bstract Suspension cultures of Coleus blumei (Lamiaceae) treated with either an elicitor preparation from the culture medium of the phytopathogenic oomycete Pythium aphanidermatum or with methyl jasmonate enhanced accumulation of rosmarinic acid approximately threefold. The specific activities of phenylalanine ammonia lyase and rosmarinic acid synthase were also enhanced after addition of the fungal elicitor. The addition of methyl jasmonate transiently increased activities of phenylalanine ammonia lyase and hydroxyphenylpyruvate reductase, whereas the activity of rosmarinic acid synthase was not stimulated and the activity of tyrosine aminotransferase was slightly and constantly enhanced. Methyl jasmonate stimulated rosmarinic acid accumulation not only when added directly to the culture medium, but also when it could reach the cells only via the gas phase. Received: 2 April 1997 / Revision received:16 June 1997 / Accepted: 15 September 1997