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1.
The tomato Leu-rich repeat receptor kinase BRASSINOSTEROID INSENSITIVE1 (BRI1) has been implicated in both peptide (systemin) and steroid (brassinosteroid [BR]) hormone perception. In an attempt to dissect these signaling pathways, we show that transgenic expression of BRI1 can restore the dwarf phenotype of the tomato curl3 (cu3) mutation. Confirmation that BRI1 is involved in BR signaling is highlighted by the lack of BR binding to microsomal fractions made from cu3 mutants and the restoration of BR responsiveness following transformation with BRI1. In addition, wound and systemin responses in the cu3 mutants are functional, as assayed by proteinase inhibitor gene induction and rapid alkalinization of culture medium. However, we observed BRI1-dependent root elongation in response to systemin in Solanum pimpinellifolium. In addition, ethylene perception is required for normal systemin responses in roots. These data taken together suggest that cu3 is not defective in systemin-induced wound signaling and that systemin perception can occur via a non-BRI1 mechanism. 相似文献
2.
Cloning the tomato curl3 gene highlights the putative dual role of the leucine-rich repeat receptor kinase tBRI1/SR160 in plant steroid hormone and peptide hormone signaling 总被引:12,自引:0,他引:12 下载免费PDF全文
Brassinosteroids (BRs) are plant steroid hormones that are essential for normal plant development. To gain better understanding of the conservation of BR signaling, the partially BR-insensitive tomato mutant altered brassinolide sensitivity1 (abs1) was identified and found to be a weak allele at the curl3 (cu3) locus. BR content is increased in both of these mutants and is associated with increased expression of DWARF: The tomato homolog of the Arabidopsis Brassinosteroid Insensitive1 Leu-rich repeat (LRR) receptor-like kinase, named tBri1, was isolated using degenerate primers. Sequence analysis of tBRI1 in the mutants cu3 and abs1 revealed that cu3 is a nonsense mutant and that abs1 is a missense mutant. A comparison of BRI1 homolog sequences highlights conserved features of BRI1 sequences, with the LRRs in close proximity to the island domain showing more conservation than N-terminal LRRs. The most homologous sequences were found in the kinase and transmembrane regions. tBRI1 (SR160) also has been isolated as the putative receptor for systemin, a plant peptide hormone. This finding suggests a possible dual role for tBRI1 in steroid hormone and peptide hormone signaling. 相似文献
3.
The plant polypeptide signal systemin induces proteinase inhibitor synthesis in tomato leaves. We show here that systemin elicits a transient depolarization of the tomato mesophyll cell membrane. Furthermore it triggers a transient decrease in the external pH of the mesophyll tissue which is followed by a sustained pH increase. In the presence of fusicoccin (which has been shown to antagonize the synthesis of proteinase inhibitors) the depolarization and transient H+ efflux are attenuated whereas the slower phase of the sustained electroneutral H+ influx persists. These results suggest that systemin-induced changes in ion transport play a role in the early phases of systemin signal transduction. 相似文献
4.
Szekeres M 《Trends in plant science》2003,8(3):102-104
Brassinosteroids, coordinating developmental events, and systemin, inducing systemic wound responses to attacks by insect pests, are newly recognized plant hormones that are perceived by plasma membrane-localized leucine-rich repeat receptor kinases. The recent characterization of the brassinosteroid receptor BRI1 from tomato revealed that this protein is identical to the previously isolated SR160 systemin receptor, strongly suggesting that both brassinosteroid and systemin signalling use the same surface receptor. 相似文献
5.
6.
A Sulfhydryl Reagent Modulates Systemic Signaling for Wound-Induced and Systemin-Induced Proteinase Inhibitor Synthesis 总被引:5,自引:0,他引:5 下载免费PDF全文
The sulfhydryl group reagent p-chloromecuribenzene sulfonic acid (PCMBS), an established inhibitor of active apoplastic phloem loading of sucrose in several plant species, is shown to be a powerful inhibitor of wound-induced and systemin-induced activation of proteinase inhibitor synthesis and accumulation in leaves of tomato plants (Lycopersicon esculentum cv Castlemart). PCMBS, supplied to young tomato plants through their cut stems, blocks accumulation of proteinase inhibitors in leaves in response to wounding. The application of systemin directly to fresh wounds enhances systemic accumulation of proteinase inhibitors to levels higher than wounding alone. Placed on fresh wounds, PCMBS severely inhibits systemic induction of proteinase inhibitors, in both the presence and absence of exogenous systemin. PCMBS inhibition can be reversed by cysteine, dithiothreitol, and glutathione. Radiolabeled systemin placed on fresh wounds is readily transported from the wounded leaves to upper leaves. However, in the presence of PCMBS, radiolabeled systemin is not transported away from wound sites. Induction of proteinase inhibitor I synthesis by oligouronides (degree of polymerization [almost equal to] 20), linolenic acid, or methyl jasmonate was not inhibited by PCMBS. The cumulative data support a possible role for sulfhydryl groups in mediating the translocation of systemin from wound sites to distal receptor sites in tomato plants and further support a role for systemin as a systemic wound signal. 相似文献
7.
Using a calmodulin (CaM) cDNA as a probe in northern analyses, transgenic tomato plants that overexpress the prosystemin gene were found to express increased levels of CaM mRNA and protein in leaves compared to wild-type plants. These transgenic plants have been reported previously to express several wound-inducible defense-related genes in the absence of wounding. Calmodulin mRNA and protein levels were found to increase in leaves of young wild-type tomato plants after wounding, or treatment with systemin, methyl jasmonate, or linolenic acid. CaM mRNA appeared within 0.5 h after wounding or supplying young tomato plants with systemin, and peaked at 1 h. The timing of CaM gene expression is similar to the expression of the wound- or systemin-induced lipoxygenase and prosystemin genes, signal pathway genes whose expression have been reported to begin at 0.5–1 h after wounding and 1–2 h earlier than the genes coding for defensive proteinase inhibitor genes. The similarities in timing between the synthesis of CaM mRNA and the mRNAs for signal pathway components suggests that CaM gene expression may be associated with the signaling cascade that activates defensive genes in response to wounding. 相似文献
8.
Sarah R Hind Robert Malinowski Roopa Yalamanchili Johannes W Stratmann 《Plant signaling & behavior》2010,5(1):42-44
Systemin is a wound signaling peptide from tomato that is important for plant defenses against herbivory. The systemin receptor was initially identified as the tomato homolog of the brassinosteroid receptor BRI1, but genetic evidence argued against this finding. However, we found that BRI1 may function as an inappropriate systemin binding protein that does not activate the systemin signaling pathway. Here we provide evidence that systemin perception is localized in a tissue-type specific manner. Mesophyll protoplasts were not sensitive to systemin, while they responded to other elicitors. We hypothesize that the elusive systemin receptor is a protein with high similarity to BRI1 which is specifically localized in vascular tissue like the systemin precursor prosystemin. Binding of systemin to BRI1 may be an artifact of transgenic BRI1-overexpressing plants, but does not take place in wild type tomato cells.Key words: systemin, systemin receptor, brassinosteroids, BRI1, BRL, protoplastsSystemin is thought to be processed from its precursor prosystemin upon insect attack and wounding of tomato leaves. Strong evidence has been gathered for an important role of (pro-)systemin in the activation of defenses against insects, and the underlying signaling pathway has been studied in detail.1 However, the perception of systemin is controversial. Meindl et al.2 and Scheer and Ryan3 identified high affinity, saturable, reversible and specific cell surface binding sites on Solanum peruvianum suspension-cultured cells which are known to be highly sensitive to systemin.4 A purification approach using a photoaffinity systemin analog identified a 160 kDa protein as the systemin receptor (SR160).5 Follow-up studies showed that overexpression of tomato 35S::SR160 in systemin-insensitive tobacco plants conferred systemin sensitivity to tobacco.6 Surprisingly, SR160 turned out to be the tomato homolog of the brassinosteroid receptor BRI1,7 which raised many questions as to the functionality of a receptor for two structurally and functionally diverse ligands. It was then shown in two independent papers that a null mutant for tomato BRI1, cu-3, exhibited a normal response to systemin.8,9 This was strong evidence that SR160/BRI1 does not represent the functional systemin receptor. Our recent data added a peculiar twist to this story. We found that overexpression of tomato BRI1 in tobacco suspension-cultured cells resulted in binding of a fluorescently labeled systemin to the plasma membranes of the transgenic tobacco cells, but not to wild type cells. Surprisingly, this did not result in BRI1-dependent signal transduction and activation of a defense response, although we detected weak BRI1-independent signaling responses to systemin.10 Together with the identification of BRI1 as the systemin receptor by Scheer and Ryan,5 the simplest explanation for this phenomenon is that BRI1 is a systemin binding protein, but not the physiological systemin receptor.Therefore and for other reasons, we suggested that the true systemin receptor may be a protein with very similar properties as BRI1, e.g., a homolog of the BRI1-like (BRL) proteins. The purification strategy employed by Scheer and Ryan5 may have resulted in binding of a photoaffinity-systemin derivative to BRI1 and one or more BRL proteins. Since BRLs and BRI1 have a very similar MW, multiple bands on a SDS-PAGE would not be detectable.Here, we would like to add another aspect of systemin perception. We provide evidence for tissue-specific systemin sensitivity and discuss how this may affect systemin binding to BRI1 and the elusive systemin receptor. Prosystemin is only present in phloem parenchyma cells.11 It can be surmised that the systemin receptor is located close to these cells. Systemin perception results in JA synthesis in companion cells of vascular bundles.12 Since JA or a JA derivative is the most likely phloem-mobile candidate for a systemic long-distance wound signal, it is thought that JA is moving from companion cells into sieve cells to reach distant parts of the plant for upregulation of wound response genes in leaf cells, including mesophyll cells.13–15Here, we tested the hypothesis that mesophyll cells lack systemin perception. We generated mesophyll protoplasts from tomato leaf material as well as protoplasts from S. peruvianum suspension-cultured cells, the same cell line that had been used for the purification of SR160/BRI1 and is known to be highly sensitive to systemin. Mesophyll protoplasts showed increased phosphorylation of MAP kinases (MPKs) in response to the elicitors flg22 and chitosan, bacterial and fungal MAMPs, respectively. However, they did not respond to systemin. In contrast, the S. peruvianum protoplasts did respond to systemin and to flg22, demonstrating that the protoplasting procedure did not compromise the systemin perception mechanism (Fig. 1). Immunocomplex kinase assays with specific antibodies against tomato MPK2 produced similar results (data not shown). Since flg22, chitosan and systemin activated the same MPKs (Fig. 1), our data indicate that systemin perception is absent in mesophyll protoplasts. Our leaf protoplasting protocol is a modification of the protocol by Yoo et al. which results in the generation of mesophyll protoplasts.16 In contrast, suspension-cultured cells do not normally represent specific cell types and it is not known why the S. peruvianum cells are highly sensitive to systemin.Open in a separate windowFigure 1Absence of systemin-induced MPK phosphorylation in mesophyll cells. Protoplasts were generated (protocol available upon request) from S. peruvianum suspension-cultured cells and from S. lycopersicum cv. MicroTom leaves. After a 1.5 hour recovery phase on ice, protoplasts were resuspended in WI medium (0.5 M mannitol, 5 mM ME S pH 5.7, 20 mM KCl), recovered for 1 hour in non-stick tubes with constant rotation on a rotary shaker at room temperature, and then treated with either water (con), 10 nM systemin (sys), 100 nM flg22, or 2.5 µg/ml chitosan (from crab shells—chi) for 10 min at room temperature. Protoplasts were analyzed for MPK phosphorylation by immunoblotting using an anti-phospho-ER K antibody (phospho-p44/42 MA PK (Erk1/2) (Thr202/Tyr204); D13.14.4E; Cell Signaling Technology) at a dilution of 1:2,000. This antibody recognizes MPKs that are phosphorylated on either the Thr and Tyr or on only the Thr within the TE Y phosphorylation motif which is conserved among plant and metazoan MPKs. It is known to recognize the tobacco MPKs SIPK and WIPK21 and Arabidopsis MPK6 and MPK3,22 the orthologs of tomato MPK1/2 and MPK3.23 Bands were visualized as described.10 Proteins on membranes were stained with Ponceau S to demonstrate equal loading.Intriguingly, BRL1, BRL2 and BRL3 are expressed in the vasculature and function in vascular pattern formation in Arabidopsis, while BRI1 is ubiquitously expressed in dividing and elongating cells. BRL3 is even specifically expressed in phloem cells.17 This matches the highly specific localization of prosystemin in the phloem parenchyma cells.11,18 The highest BRI1 expression is found in growing parts of young leaves17,19 while prosystemin is specifically present in the phloem parenchyma cells throughout all developmental stages.11 In this context, it is also interesting to note that application of systemin to tomato plants via the cut stem results in rapid and strong MPK activation. In this assay, systemin is delivered to leaf cells via the transpiration stream and therefore present in vascular tissue.20Based on the combined evidence, we propose that the true systemin receptor is a BRL or similar protein which is expressed in phloem cells in the vicinity of the parenchyma cells that express prosystemin, but not in mesophyll cells. Because of the similarity between BRLs and BRI1, BRI1 was erroneously identified as the systemin receptor. Inappropriate binding of systemin to BRI1 is consistent with the high similarity between BRI1 and BRLs. However, because of the tissue-specificity of the systemin signaling pathway, inappropriate binding of systemin to BRI1 may rarely occur in wild type plants and may not pose an interference problem for either systemin or brassinosteroid signaling. 相似文献
9.
Autoradiographic and biochemical evidence for the systemic translocation of systemin in tomato plants 总被引:9,自引:0,他引:9
Javier Narváez-Vásquez Gregory Pearce Martha L. Orozco-Cardenas Vincent R. Franceschi Clarence A. Ryan 《Planta》1995,195(4):593-600
The movement of systemin, the 18-amino-acid polypeptide inducer of proteinase inhibitors in tomato (Lycopersicon esculentum L.) plants, was investigated in young tomato plants following the application of [14C]systemin to wounds on the surface of leaves. Wholeleaf autoradiographic analyses revealed that [14C]systemin was distributed throughout the wounded leaf within 30 min, and then during the next several hours was transported to the petiole, to the main stem, and to the upper leaves. The movement of [14C]systemin was similar to the movement of [14C]sucrose when applied to leaf wounds, except that sucrose was slightly more mobile than systemin. Analyses of the radioactivity in the petiole phloem exudates at intervals over a 5-h period following the application of [14C]systemin to a wound demonstrated that intact [14C]systemin was present in the phloem over the entire time, indicating that the polypeptide was either stable for long periods in the phloem or was being continually loaded into the phloem from the source leaf. The translocation pathway of systemin was also investigated at the cellular level, using light microscopy and autoradiography. Within 15 min after application of [3H]systemin to a wound on a terminal leaflet, it was found distributed throughout the wounded leaf and was primarily concentrated in the xylem and phloem tissues within the leaf veins. After 30 min, the radioactivity was found mainly associated with vascular strands of phloem tissue in the petiole and, at 90 min, label was found in the phloem of the main stem. Altogether, these and previous results support a role for systemin as a systemic wound signal in tomato plants.The authors acknowledge the Washington State University Electron Microscope Center and staff for their technical advice and collaboration. We also thank Greg Wichelns for growing our plants and Dr. Steven Doares for providing [3H]systemin. This research was supported in part by the Washington State College of Agriculture and Home Economics Project No. 1791 and National Science Foundation grants IBN 9117795 and IBN 9104542 相似文献
10.
The tomato suppressor of prosystemin-mediated responses2 gene encodes a fatty acid desaturase required for the biosynthesis of jasmonic acid and the production of a systemic wound signal for defense gene expression 总被引:9,自引:0,他引:9 下载免费PDF全文
Genetic analysis of the wound response pathway in tomato indicates that systemin and its precursor protein, prosystemin, are upstream components of a defensive signaling cascade that involves the synthesis and subsequent action of the octadecatrienoic acid (18:3)-derived plant hormone jasmonic acid (JA). The suppressor of prosystemin-mediated responses2 (spr2) mutation, which was isolated previously as a suppressor of (pro)systemin-mediated signaling, impairs wound-induced JA biosynthesis and the production of a long-distance signal for the expression of defensive Proteinase inhibitor genes. Using a map-based cloning approach, we demonstrate here that Spr2 encodes a chloroplast fatty acid desaturase involved in JA biosynthesis. Loss of Spr2 function reduced the 18:3 content of leaves to <10% of wild-type levels, abolished the accumulation of hexadecatrienoic acid, and caused a corresponding increase in the level of dienoic fatty acids. The effect of spr2 on the fatty acyl content of various classes of glycerolipids indicated that the Spr2 gene product catalyzes most, if not all, omega3 fatty acid desaturation within the "prokaryotic pathway" for lipid synthesis in tomato leaves. Despite the reduced levels of trienoic fatty acids, spr2 plants exhibited normal growth, development, and reproduction. However, the mutant was compromised in defense against attack by tobacco hornworm larvae. These results indicate that jasmonate synthesis from chloroplast pools of 18:3 is required for wound- and systemin-induced defense responses and support a role for systemin in the production of a transmissible signal that is derived from the octadecanoid pathway. 相似文献
11.
Small signaling molecules that mediate cell-cell communication are essential for developmental regulation in multicellular organisms. Among them are the steroids and peptide hormones that regulate growth in both plants and animals. In plants, brassinosteroids (BRs) are perceived by the cell surface receptor kinase BRI1, which is distinct from the animal steroid receptors. Identification of components of the BR signaling pathway has revealed similarities to other animal and plant signal transduction pathways. Recent studies demonstrated that tomato BRI1 (tBRI1) perceives both BR and the peptide hormone systemin, raising new questions about the molecular mechanism and evolution of receptor-ligand specificity. 相似文献
12.
Suppressors of systemin signaling identify genes in the tomato wound response pathway. 总被引:17,自引:0,他引:17
In tomato plants, systemic induction of defense genes in response to herbivory or mechanical wounding is regulated by an 18-amino-acid peptide signal called systemin. Transgenic plants that overexpress prosystemin, the systemin precursor, from a 35S::prosystemin (35S::prosys) transgene exhibit constitutive expression of wound-inducible defense proteins including proteinase inhibitors and polyphenol oxidase. To study further the role of (pro)systemin in the wound response pathway, we isolated and characterized mutations that suppress 35S::prosys-mediated phenotypes. Ten recessive, extragenic suppressors were identified. Two of these define new alleles of def-1, a previously identified mutation that blocks both wound- and systemin-induced gene expression and renders plants susceptible to herbivory. The remaining mutants defined four loci designated Spr-1, Spr-2, Spr-3, and Spr-4 (for Suppressed in 35S::prosystemin-mediated responses). spr-3 and spr-4 mutants were not significantly affected in their response to either systemin or mechanical wounding. In contrast, spr-1 and spr-2 plants lacked systemic wound responses and were insensitive to systemin. These results confirm the function of (pro)systemin in the transduction of systemic wound signals and further establish that wounding, systemin, and 35S::prosys induce defensive gene expression through a common signaling pathway defined by at least three genes (Def-1, Spr-1, and Spr-2). 相似文献
13.
Nicholas Holton Kate Harrison Takao Yokota Gerard J Bishop 《Plant signaling & behavior》2008,3(1):54-55
Brassinosteroids (BRs) are perceived by Brassinosteroid Insensitive 1 (BRI1), that encodes a leucine-rich repeat receptor kinase. Tomato BRI1 has previously been implicated in both systemin and BR signalling. The role of tomato BRI1 in BR signalling was confirmed, however it was found not to be essential for systemin/wound signalling. Tomato roots were shown to respond to systemin but this response varied according to the species and growth conditions. Overall the data indicates that mutants defective in tomato BRI1 are not defective in systemin-induced wound signalling and that systemin perception can occur via a non-BRI1 mechanism.Key words: tomato BRI1, brassinosteroids, systemin, wound signallingBrassinosteroids (BRs) are steroid hormones that are essential for normal plant growth. The most important BR receptor in Arabidopsis is BRASSINOSTERIOD INSENSITIVE 1 (BRI1), a serine/threonine kinase with a predicted extracellular domain of ∼24 leucine-rich repeats (LRRs).1,2 BRs bind to BRI1 via a steroid-binding domain that includes LRR 21 and a so-called “island” domain.2,3 In tomato a BRI1 orthologue has been identified that when mutated, as in the curl3 (cu3) mutation, results in BR-insensitive dwarf plants.4 Tomato BRI1 has also been purified as a systemin-binding protein.5 Systemin is an eighteen amino acid peptide, which is produced by post-translational cleavage of prosystemin. Systemin has been implicated in wound signalling and is able to induce the production of jasmonate, protease inhibitors (PIN) and rapid alkalinization of cell suspensions (reviewed in ref. 6).To clarify whether tomato BRI1 was indeed a dual receptor it was important to first confirm its role in BR signalling. Initially this was carried out by genetic complementation of the cu3 mutant phenotype.7 Overexpression of tomato BRI1 restored the dwarf phenotype and BR sensitivity and normalized BR levels (35S:TomatoBRI1 complemented line Wt* cu3* 6-deoxocathasterone 566 964 676 6-deoxoteasterone nd 47 48 3-dehydro-6-deoxoteasterone 87 62 69 6-deoxotyphasterol nd 588 422 6-deoxocastasterone 1,755 6,247 26,210 castasterone 255 637 17,428 brassinolide nd nd nd