Molecular details of tomato extensin and glycine-rich protein gene expression

In a recent publication (Plant Molecular Biology 16: 547–565 (1991)) Showalter et al. described the isolation and initial characterization of fifteen extensin and extensin-like tomato cDNAs. These cDNAs were determined to fall into five distinct classes; class I and II clones encoded extensins, class III and V clones encoded glycine-rich proteins (GRPs), and class IV clones encoded a portion of a GRP sequence on one DNA strand and a portion of an extensin sequence on the other DNA strand. In this publication, a more detailed analysis of the expression of these cDNA classes was performed with respect to wounding in various tomato organs, development, kinetics and systemic extent of the wound response, ethylene treatment, abscisic acid (ABA) treatment, and drought stress by using RNA gel blot hybridizations. In general, extensin gene expression was readily detected in stems and roots, but not in leaves. With both class I and II extensin cDNA probes, wound-induced accumulation of mRNA in stems was first detected between 4 and 8 h after wounding with maximal accumulation occurring after 12 h. Moreover, these extensin wound responses were detected locally at the wound site but not systemically. Expression of the class III GRP was largely limited to wounded stem tissue. Initial detection and maximal accumulation of the class III GRP mRNA was similar to the extensins mRNAs; however, this GRP wound response occurred both locally and systemically. Additionally, abscisic acid treatment and drought stress resulted in the marked accumulation of the class III GRP mRNA in tomato stems, but did not alter the expression of the other cDNA classes. In contrast, expression of the class V GRP occurred in stems and roots and to a lesser extent in leaves and decreased in response to wounding over a 24 h time period. The class V GRP wound response was further characterized by an early, transient accumulation of mRNA occurring 2–4 h after wounding in stems and by its local nature.     

Wounding changes the spatial expression pattern of the arabidopsis plastid omega-3 fatty acid desaturase gene (FAD7) through different signal transduction pathways.

The Arabidopsis FAD7 gene encodes a plastid omega-3 fatty acid desaturase that catalyzes the desaturation of dienoic fatty acids in membrane lipids. The mRNA levels of the Arabidopsis FAD7 gene in rosette leaves rose rapidly after local wounding treatments. Wounding also induced the expression of the FAD7 gene in roots. To study wound-responsive expression of the FAD7 gene in further detail, we analyzed transgenic tobacco plants carrying the -825 Arabidopsis FAD7 promoter-beta-glucuronidase fusion gene. In unwounded transformants, FAD7 promoter activity was restricted to the tissues whose cells contained chloroplasts. Activation of the FAD7 promoter by local wounding treatments was more substantial in stems (29-fold) and roots (10-fold) of transgenic plants than it was in leaves (approximately two-fold). Significant induction by wounding was observed in the overall tissues of stems and included trichomes, the epidermis, cortex, vascular system, and the pith of the parenchyma. Strong promoter activity was found preferentially in the vascular tissues of wounded roots. These results indicate that wounding changes the spatial expression pattern of the FAD7 gene. Inhibitors of the octadecanoid pathway, salicylic acid and n-propyl gallate, strongly suppressed the wound activation of the FAD7 promoter in roots but not in leaves or stems. In unwounded plants, exogenously applied methyl jasmonate activated the FAD7 promoter in roots, whereas it repressed FAD7 promoter activity in leaves. Taken together, wound-responsive expression of the FAD7 gene in roots is thought to be mediated via the octadecanoid pathway, whereas in leaves, jasmonate-independent wound signals may induce the activation of the FAD7 gene. These observations indicate that wound-responsive expression of the FAD7 gene in aerial and subterranean parts of plants is brought about by way of different signal transduction pathways.     

Fast upward propagation of the wound signal that systemically elevates phosphatidic acid

Phosphatidic acid (PA) increases in response to wounding at the neighboring unwounded leaf as well as at the wounded leaf of many plants (Lee et al., 1997). This indicates that a signal propagates from the wounded leaf to its neighboring leaves. In this paper, we report the speed and direction of propagation for a systemic wound signal that elevates PA. When a leaf of a soybean (Glycine max) seedling at the 2-leaf-stage was wounded, the PA level of the neighboring leaf did not change within the first min, but did increase significantly in 2 min, returning to the control level after 15 min. This implies that the systemic wound signal was generated at least within 2 min of wounding, and was propagated at a speed of at least 10–16 mm/min. When we wounded individual leaves of soybean and tobacco (Nicotiana tabaccum) seedlings that had 3 or 4 leaves, PA levels were elevated only in the younger leaves located above the wounded leaf, but not in the older, lower leaves. Thus, the PA-elevating wound signal preferentially moves upward in these plants.     

Wounding of Arabidopsis leaves causes a powerful but transient protection against Botrytis infection

Physical injury inflicted on living tissue makes it vulnerable to invasion by pathogens. Wounding of Arabidopsis thaliana leaves, however, does not conform to this concept and leads to immunity to Botrytis cinerea , the causal agent of grey mould. In wounded leaves, hyphal growth was strongly inhibited compared to unwounded controls. Wound-induced resistance was not associated with salicylic acid-, jasmonic acid- or ethylene-dependent defence responses. The phytoalexin camalexin was found to be involved in this defence response as camalexin-deficient mutants were not protected after wounding and the B. cinerea strains used here were sensitive to this compound. Wounding alone did not lead to camalexin production but primed its accumulation after inoculation with B. cinerea , further supporting the role of camalexin in wound-induced resistance. In parallel with increased camalexin production, genes involved in the biosynthesis of camalexin were induced faster in wounded and infected plants in comparison with unwounded and infected plants. Glutathione was also found to be required for resistance, as mutants deficient in γ-glutamylcysteine synthetase showed susceptibility to B. cinerea after wounding, indicating that wild-type basal levels of glutathione are required for the wound-induced resistance. Furthermore, expression of the gene encoding glutathione- S -transferase 1 was primed by wounding in leaves inoculated with B. cinerea . In addition, the priming of MAP kinase activity was observed after inoculation of wounded leaves with B . cinerea compared to unwounded inoculated controls. Our results demonstrate how abiotic stress can induce immunity to virulent strains of B. cinerea , a process that involves camalexin and glutathione.     

Wound-induced and developmental activation of a poplar tree chitinase gene promoter in transgenic tobacco

Wounding hybrid poplar (Populus trichocarpa × P. deltoides) trees results in the expression of novel wound-inducible (win) mRNAs thought to encode proteins involved in defense against pests and pathogens. Members of thewin6 gene family encode acidic multi-domain chitinases, with combined structure and charge characteristics that differ from previously described chitinases.Win6 expression has been shown to occur in pooled unwounded leaves of a wounded (on multiple leaves) poplar plant. Here we demonstrate that wounding a single leaf induceswin6 expression locally, in the wounded leaf, and remotely, in specific unwounded leaves with strong vascular connections to the wounded leaf. We also demonstrate that awin6 promoter--glucuronidase (GUS) gene fusion (win6-GUS) responds to wounding locally and remotely in transgenic tobacco. These data indicate that the poplarwin6 promoter has regulatory elements that are responsive to wound signals in the heterologous host. In addition,win6-GUS is developmentally activated in unwounded young leaves and floral tissues of transgenic tobacco. Similar developmental expression patterns are found to occur forwin6 in poplar trees, demonstrating that a herbaceous plant can serve as a host for woody tree transgene analysis and can accurately predict expression patterns in tree tissues (e.g. flowers) that would be difficult to study in free-living trees.     

Arabidopsis thaliana Atvsp is homologous to soybean VspA and VspB,genes encoding vegetative storage protein acid phosphatases,and is regulated similarly by methyl jasmonate,wounding, sugars,light and phosphate

The soybean vegetative storage proteins, VSP and VSP, are acid phosphatases that accumulate to very high levels in hypocotyls, young leaves and flowers and pods. The genes encoding the soybean VSP are activated by jasmonate, wounding, sugars and light and down regulated by phosphate and auxin. In this study, expression of an Arabidopsis thaliana gene (Atvsp) encoding a protein homologous to soybean Vsp and Vsp, was examined and compared to expression of the soybean Vsp genes. Atvsp mRNA was present at high levels in flowers and buds and at low levels in roots, stems, leaves and siliques. Expression of Atvsp in leaves could be induced by wounding or by treatment of illuminated plants with methyl jasmonate and sucrose. Roots of plants with wounded leaves also accumulated Atvsp mRNA indicating that this gene can be regulated by a transmissible wound signal. Phosphate partially inhibited expression of Atvsp. Arabidopsis proteins of 29 and 30 kDa crossreacted with antibodies against soybean VSP. These proteins were very abundant in flowers and the proteins accumulated in leaves and roots of plants treated with methyl jasmonate. The level of these proteins in flowers was similar to the levels of soybean VSP in young soybean leaves. Overall, these data indicate that Arabidopsis Atvsp and soybean VspA/B genes are regulated similarly and that in both plants, the gene products can accumulate to high levels. This suggests that genes homologous to VspA/B may be of greater general significance than previously recognized.     

Local and systemic changes in arginine decarboxylase activity, putrescine levels and putrescine catabolism in wounded oilseed rape

* Here we report the effect of mechanical wounding on putrescine biosynthesis and catabolism in oilseed rape (Brassica napus ssp. oleifera). * The lamina of first leaves was wounded by crushing with forceps, and first and second leaves were harvested at various intervals over a 24 h period. Levels of free polyamines were measured and activities of enzymes of polyamine biosynthesis and catabolism were assayed in the harvested tissue. * Mechanical wounding of the first leaves led to significant, but transient, increases in arginine decarboxylase (ADC) activity and levels of free putrescine in the wounded first leaf and in unwounded second leaves. The increased putrescine appeared to be the result of a combination of increased ADC activity, coupled with reduced putrescine catabolism, as activity of the oxidative enzyme diamine oxidase was significantly reduced following wounding, both locally and systemically. * The role of the increased free putrescine in the wound response of oilseed rape is not known, although the possibility that it is used to form putrescine conjugates is worthy of further investigation.     

Rapid accumulation of trihydroxy oxylipins and resistance to the bean rust pathogen Uromyces fabae following wounding in Vicia faba

BACKGROUND AND AIMS: Insect damage to plants leads to wound-activated responses directed to healing of damaged tissues, as well as activation of defences to prevent further insect damage. Negative cross-talk exists between the jasmonic acid-based signalling system that is activated upon insect attack and the salicylic acid-based system frequently activated following pathogen infection. Thus, insect attack may compromise the ability of the plant to defend itself against pathogens and vice versa. However, insect herbivory and mechanical wounding have been shown to reduce fungal infections on some plants, although the underlying mechanisms remain to be defined. This work examines the effects of mechanical wounding on rust infection both locally and systemically in the broad bean, Vicia faba and follows changes in oxylipins in wounded leaves and unwounded leaves on wounded plants. METHODS: The lamina of first leaves was wounded by crushing with forceps, and first and second leaves were then inoculated, separately, with the rust Uromyces fabae at various times over a 24 h period. Wounded first leaves and unwounded second leaves were harvested at intervals over a 24 h period and used for analysis of oxylipin profiles. KEY RESULTS Mechanical wounding of first leaves of broad bean led to significantly reduced rust infection in the wounded first leaf as well as the unwounded second leaf. Increased resistance to infection was induced in plants inoculated with rust just 1 h after wounding and was accompanied by rapid and significant accumulation of jasmonic acid and two trihydroxy oxylipins in both wounded first leaves and unwounded second leaves. The two trihydroxy oxylipins were found to possess antifungal properties, reducing germination of rust spores. CONCLUSIONS: These results demonstrate the rapidity with which resistance to pathogen infection can be induced following wounding and provides a possible mechanism by which pathogen infection might be halted.