New Clothes for the Jasmonic Acid Receptor COI1: Delayed Abscission,Meristem Arrest and Apical Dominance

In a screen for delayed floral organ abscission in Arabidopsis, we have identified a novel mutant of CORONATINE INSENSITIVE 1 (COI1), the F-box protein that has been shown to be the jasmonic acid (JA) co-receptor. While JA has been shown to have an important role in senescence, root development, pollen dehiscence and defense responses, there has been little focus on its critical role in floral organ abscission. Abscission, or the detachment of organs from the main body of a plant, is an essential process during plant development and a unique type of cell separation regulated by endogenous and exogenous signals. Previous studies have indicated that auxin and ethylene are major plant hormones regulating abscission; and here we show that regulation of floral organ abscission is also controlled by jasmonic acid in Arabidopsis thaliana. Our characterization of coi1-1 and a novel allele (coi1-37) has also revealed an essential role in apical dominance and floral meristem arrest. In this study we provide genetic evidence indicating that delayed abscission 4 (dab4-1) is allelic to coi1-1 and that meristem arrest and apical dominance appear to be evolutionarily divergent functions for COI1 that are governed in an ecotype-dependent manner. Further characterizations of ethylene and JA responses of dab4-1/coi1-37 also provide new information suggesting separate pathways for ethylene and JA that control both floral organ abscission and hypocotyl growth in young seedlings. Our study opens the door revealing new roles for JA and its interaction with other hormones during plant development.     

Characterization of a Non-abscission Mutant in Lupinus angustifolius L.: Physiological Aspects

A single-gene recessive mutant (Abs^-) of Lupinus angustifoliusL. ‘Danja’ that does not abscise any organs wascompared with its parent during continuous exposure of explantsfrom 14 d old seedlings to 10 µl l^-1ethylene. Both endo-(1,4)-ß- D -glucanase (cellulase) and polygalacturonase(PGA) activities increased significantly and progressively inpetiole-stem abscission zones of the parent before the onsetof abscission, and were reflected in a rapid decline in breakstrengthfrom 300 to 70 g within 32 h. In the mutant there was negligibleincrease in hydrolytic enzyme activity, breakstrength declinedslowly (to 180–200 g by 72 h) and there was no abscission.Isoelectric focusing showed two cellulase isoforms (pI 5.0 andpI 8.5) expressed in abscission zones of the parent; these wereexpressed at much lower levels in the mutant. These data areinterpreted to indicate that expression of at least two formsof cellulase activity is enhanced by ethylene in normal petioleabscission zones of lupin. PGA activity also increased in theabscission zone tissue of the parent but to a lesser extentin that of the mutant. We attribute the Abs^-phenotype to mutationof a gene regulating ethylene-responsive expression of abscission-specifichydrolytic enzymes. Copyright 2001 Annals of Botany Company Lupinus angustifolius, abscission, breakstrength, cellulase, ethylene, legume, lupin, mutant, polygalacturonase     

Hormonal control of floral abscission in zucchini squash (<Emphasis Type="Italic">Cucurbita pepo</Emphasis>)

In the zucchini squash, Cucurbita pepo, a well coordinated abscission of the female flower during fruit set is essential to obtain a fruit of commercial value. In Spain zucchini is mainly produced in greenhouses in Almería, where high temperatures during the spring-summer period provoke a cultivar-dependent defect in fruits known as the “sticky flower” syndrome. This disorder is characterised by an arrest in growth and maturation of floral organs, and a lack of female floral abscission, thus diminishing fruit shelf-life, commercial quality and value. The aim of the present work was to improve knowledge of the abscission process in C. pepo to better understand the fundamental causes of this disorder. The anatomical analysis of abscission shows a well defined male floral abscission zone (AZ), few hours after anthesis, which differs from the female zone which is not differentiated from the adjacent tissue until the abscission process has begun, and which occurs as a consequence of AZ cell enlargement and the dissolution of their cell walls. To evaluate the role of ethylene and auxins in the regulation of floral abscission in zucchini we performed several treatments, with: ethylene, added as 0.25% ethrel solution; AVG, the inhibitor of ethylene synthesis, at 100 μM; indol-3-acetic acid, 100 μM; and TIBA, the inhibitor of auxin polar transport, at 10 mM. These treatments show that ethylene is an accelerator of zucchini floral abscission, and also promotes abscission in isolated AZs of sticky flowers. On the other hand, IAA delays abscission of the female flowers, whilst the inhibitor of auxin polar transport promotes it. The activity of the cell wall hydrolytic enzymes, polygalacturonase and cellulase, sharply increased just before the shedding of zucchini floral organs (72 h after anthesis). Moreover, both enzyme activities were induced by ethylene, which partly explains the ethylene promoting effect.     

Hawaiian skirt: an F-box gene that regulates organ fusion and growth in Arabidopsis

A fast neutron-mutagenized population of Arabidopsis (Arabidopsis thaliana) Columbia-0 wild-type plants was screened for floral phenotypes and a novel mutant, termed hawaiian skirt (hws), was identified that failed to shed its reproductive organs. The mutation is the consequence of a 28 bp deletion that introduces a premature amber termination codon into the open reading frame of a putative F-box protein (At3g61590). The most striking anatomical characteristic of hws plants is seen in flowers where individual sepals are fused along the lower part of their margins. Crossing of the abscission marker, Pro(PGAZAT):beta-glucuronidase, into the mutant reveals that while floral organs are retained it is not the consequence of a failure of abscission zone cells to differentiate. Anatomical analysis indicates that the fusion of sepal margins precludes shedding even though abscission, albeit delayed, does occur. Spatial and temporal characterization, using Pro(HWS):beta-glucuronidase or Pro(HWS):green fluorescent protein fusions, has identified HWS expression to be restricted to the stele and lateral root cap, cotyledonary margins, tip of the stigma, pollen, abscission zones, and developing seeds. Comparative phenotypic analyses performed on the hws mutant, Columbia-0 wild type, and Pro(35S):HWS ectopically expressing lines has revealed that loss of HWS results in greater growth of both aerial and below-ground organs while overexpressing the gene brings about a converse effect. These observations are consistent with HWS playing an important role in regulating plant growth and development.     

Characterization of a non-abscission mutant in Lupinus angustifolius. I. Genetic and structural aspects

A spontaneous mutant, Abs, that does not abscise any organs despite an apparently normal pattern of growth and senescence was isolated from among plants of Lupinus angustifolius cv. 'Danja'. Abs was found to be a recessive single gene mutation, and it was proposed that the gene for the original mutant phenotype, referred to as Abs, be designated abs1. An artificially induced mutant allelic to abs1 was also obtained and a non-allelic mutant phenotype, Delabs (delayed abscission), which was designated abs2. Morphological and cytological features of the abscission process under conditions of natural and ethylene-induced senescence were compared in the wild-type parent and Abs mutant. In the parent genotype abscission under natural conditions is similar to many other species, consisting of a stage of cell division forming an abscission zone, activation of the cytoplasm of zone cells, dissolution of the middle lamella, disorganization of fibrillar wall structure, and cell separation. A slightly different pattern of abscission zone development was observed for ethylene-treated explants of the parent, mainly with respect to features of cell division and cell enlargement. In Abs no abscission occurred for any abscission sites under conditions of natural senescence or with ethylene treatment of small shoot explants. However, relatively normal abscission zones were differentiated at all sites in the mutant except that extensive cell wall disorganization did not occur. Ethylene production by leaves or other organs of the mutant was no different from that of Danja. Application of copper salts or hydrogen peroxide, droughting, waterlogging, or application of abscisic acid (ABA) increased ethylene production equally in both genotypes but did not result in abscission in the mutant. Release of root cap border cells, the only other cell separation process examined, was similar in each genotype. The study concludes that the mutation is quite specific to the abscission process and may be due to a lack of or delay in the expression of hydrolytic enzyme(s) associated specifically with abscission zone differentiation and separation.     

Abscission of Azolla branches induced by ethylene and sodium azide

Treatment with ethylene accelerated the abscission of branches of Azolla filiculoides plants. An Azolla plantlet treated with ethylene at 10 microl liter(-1) divided into 4-5 fragments after a lag period of 6-8 h. Ethylene-induced abscission was effectively inhibited by cycloheximide and was associated with an increase in the activities of cellulase and polygalacturonase. At the fracture surface abscised after treatment with ethylene, dissolution of the primary walls of the abscission zone cells was apparent. However, the middle lamella between abscission zone cells was still present. Immunoelectron microscopy using anti-unesterified pectin (JIM5) and anti-methylesterified pectin (JIM7) monoclonal antibodies revealed the presence of both JIM5 and JIM7 epitopes in the wall between abscission zone cells of branches before abscission occurred. In the middle lamella remaining after ethylene-induced abscission, only JIM7 epitopes were observed. The features of ethylene-induced abscission described herein were different from those of the rapid abscission induced by sodium azide, which implies that they are mediated by different mechanisms. The possible mechanisms are discussed.     

Perianth Abscission in Montbretia (Crocosmia x crocosmiiflora)

The anatomy, histochemistry, kinetics and hormonal regulationof perianth abscission in Crocosmia x crocosmiiflora (Montbretia)has been investigated. The abscission zone is anatomically welldefined, with cell divisions occurring in this region at anthesis.Abscission is first detectable 3 d after perianth opening, whenthe walls of a group of cells beneath the adaxial epidermisshow reduced staining with polyanion-specific stains, and adecline in penanth break strength also occurs. Abscission isachieved by cell wall breakage in thc abscission zone, whichprogresses eccentrically from the adaxial epidermis throughthe abscission zone, rather than the separation of intact cellsas occurs in flowers of dicotyledons. Experiments on detachedflowers suggest similarities in the hormonal regulation of abscissionin Crocosmia to that of dicotyledons, in that an ethylene promotion,and possibly an auxin inhibition, mechanism may exist in Crocosmia.Ovary expansion occurs throughout the development and senescenceof unpollinated flowers, but does not appear to be the solecause of wall breakage in the abscission zone. It is suggestedthat hormonally regulated wall hydrolases weaken the cell wallsin the abscission zone, and allow wall breakage and subsequentabscission to occur. Cdrocosmia x crocosmiiflora, Montbretia, anatomy, breakstrength, cell wall changes, histochemistry, flowers, monocotyledons, perianth, senescence, ethylene, auxin     

Lack of ethylene involvement in tulip tepal abscission

The tepals of cut flowers of Tulipa hybrida cv. Golden Apeldoorn and Tulipa kaufmanniana cv. Shakespeare abscise 3–4 days after harvest. The weakening of the abscission zones is accompanied by cell wall breakdown and the separation of 3–4 rows of intact cells at the base of the tepal. During senescence, there is no ethylene climacteric and ethylene production rates remain low, between 0.07 and 0.4 nl g⁻¹ fresh weight h⁻¹. Adding 3–5 μl l⁻¹ ethylene slightly accelerated the weakening of the abscission zones but had no effect on the time of first abscission. Neither 0.5 m M silver thiosulphate nor 5 m M aminoethoxyvinylglycine delayed the time to abscission. It is concluded that tulip tepal fall does not involve primary regulation by ethylene, unlike the majority of other abscission systems that have been studied.     

Cellulase activity and localization during induced abscission of Coleus blumei

Treatment with dimethipin (2,3-dihydro-5,6-dimethyl-1,4-dithiin 1,1,4,4 tetroxide) inhibited the increase in cellulase activity and decrease in breakstrength associated with the normal course of abscission in Coleus. Application of the surfactant UBI-1126 (Emery OAL 20 in isopropyl alcohol) increased cellulase activity and accelerated the process of abscission in Coleus expiants within 24 h of application. Cellulase activity was localized histochemically at the electron microscopic level in surfactant-treated tissue. The enzyme activity was localized primarily in the cell wall, middle lamella, and paramural bodies of abscission zone cells.     

Compounds Interacting with the Ethylene Receptor in Plants

Abstract: Some of the compounds binding to the ethylene receptor induce an ethylene response, but others prevent it. The compounds preventing an ethylene response have been developed into a means for protecting plants against ethylene and extending the life of some plant material. 1-Methylcyclopropene (1-MCP), a compound now commercially available under the names EthylBloc and SmartFresh™, is currently being used on flowers, fruit and vegetables with great success. In ethylene sensitive flowers, among other responses, it prevents senescence and abscission of plant organs; in fruit and vegetables it slows down the ripening process. Other similar compounds are now being developed for a range of methods of application.     

The MADS box gene, FOREVER YOUNG FLOWER, acts as a repressor controlling floral organ senescence and abscission in Arabidopsis

The ectopic expression of a MADS box gene FOREVER YOUNG FLOWER (FYF) caused a significant delay of senescence and a deficiency of abscission in flowers of transgenic Arabidopsis. The defect in floral abscission was found to be due to a deficiency in the timing of cell separation of the abscission zone cells. Down-regulation of INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) may contribute to the delay of the floral abscission in 35S:FYF flowers. FYF was found to be highly expressed in young flowers prior to pollination and was significantly decreased after pollination, a pattern that correlated with its function. Ethylene insensitivity in senescence/abscission and the down-regulation of ETHYLENE RESPONSE DNA-BINDING FACTOR 1 (EDF1) and EDF2, downstream genes in the ethylene response, in 35S:FYF Arabidopsis suggested a role for FYF in regulating senescence/abscission by suppressing the ethylene response. This role was further supported by the fact that 35S:FYF enhanced the delay of flower senescence/abscission in ethylene response 1 (etr1), ethylene-insensitive 2 (ein2) and constitutive triple response 1 (ctr1) mutants, which have defects in upstream genes of the ethylene signaling pathway. The presence of a repressor domain in the C-terminus of FYF and the enhancement of the delay of senescence/abscission in FYF+SRDX (containing a suppression motif) transgenic plants suggested that FYF acts as a repressor. Indeed, in FYF-DR+VP16 transgenic dominant-negative mutant plants, in which FYF was converted to a potent activator by fusion to a VP16-AD motif, the senescence/abscission of the flower organs was significantly promoted, and the expression of BOP2, IDA and EDF1/2 was up-regulated. Our data suggest a role for FYF in controlling floral senescence/abscission by repressing ethylene responses and regulating the expression of BOP2 and IDA in Arabidopsis.     

Effect of octanoic acid on ethylene-mediated processes in Arabidopsis

We have examined whether octanoic acid (OA) one of the short chain saturated fatty acids (SCSFA), increases ethylene response in the following three ethylene-mediated processes: a) hypocotyl growth in darkness; b) formation of new flowers; c) flower abscission. These processes were examined in the presence or absence of exogenous ethylene in Arabidopsis wild type (WT) and in the ethylene-insensitive mutants, etr1-3 and ein2-1 and in the ethylene over-producer mutant eto1-1. Our results show that OA decreased hypocotyl length of WT in the absence or presence of exogenous ethylene, apparently showing that OA acts via augmentation of ethylene action. However, the hypocotyl growth inhibition could not be ascribed to increased ethylene sensitivity since application of inhibitors of ethylene synthesis (aminoethoxyvinylglycine; AVG) or action (1-methylcyclopropene;1-MCP) to WT seedlings did not prevent specifically the OA-induced growth inhibition. Also, OA inhibited hypocotyl growth in the mutants etr1-3 and ein2-1 in a similar pattern to that obtained in WT. On the other hand, OA had no effect on flower formation neither in WT, etr1-3 and eto1-1, in which ethylene reduced flower formation, nor in the ein2-1 mutant, in which ethylene had no effect. OA also did not increase flower abscission in WT or in the mutants etr1-3 and ein2-1 neither in the absence nor in the presence of ethylene. However, OA has augmented flower abscission in the mutant eto1-1 only in the absence of exogenous ethylene. This result might indicate that the effect of OA on eto1-1 is specific to this mutant and is not due to general deleterious effects inflicted by OA. Taken together, our results show that in general OA does not augment ethylene response in Arabidopsis, but it might affect ethylene action in flower abscission of the ethylene-overproducer mutant.     

Role of Ethylene Biosynthesis and Auxin Content and Transport in High Temperature-Induced Abscission of Pepper Reproductive Organs

High temperatures induced abscission of pepper (Capsicum annuum L. cv. Maor) reproductive organs at various developmental stages. The role of ethylene biosynthesis and auxin economy in high temperature-induced abscission is described. High temperatures somewhat increased ethylene production in the reproductive organs, but the highest temperature treatment, which was the most active in inducing reproductive organ abscission, decreased it. In contrast to ethylene, 1-aminocyclopropane-1-carboxylic acid levels increased significantly in response to high temperatures and correlated positively with the increase in temperature. High temperatures reduced indole-3-acetic acid levels and particularly auxin transport capacity in the reproductive organs. The data suggest that the reduction of auxin transport capacity is the major mechanism by which high temperatures induce reproductive organ abscission in pepper. Received September 27, 1996; accepted March 13, 1997     

Ethylene and the regulation of senescence processes in transgenic Nicotiana sylvestris plants

BACKGROUND AND AIMS: Exposure of plants to ethylene can influence a spectrum of developmental processes including organ senescence and abscission. The aim of this study was to examine the role of the gaseous regulator in Nicotiana sylvestris plants exhibiting a silenced or constitutive ethylene response. METHODS: Transgenic N. sylvestris plants were generated that either ectopically expressed the Arabidopsis mutant ethylene receptor ETR1-1 or the tomato EIN3-like (LeEIL1) gene. Highly expressing homozygous lines were selected and the time-course of development, from germination to organ senescence, was studied. KEY RESULTS: Fifty percent of the homozygous Pro(35S):ETR1-1 lines examined showed a high susceptibility to collapse prior to flowering, with plant death occurring within a few days of leaf wilting. The time-course of leaf senescence in the remaining Pro(35S):ETR1-1 lines was visibly arrested compared to wild type (negative segregant) plants and this observation was reaffirmed by chlorophyll and protein analysis. Petal necrosis was also delayed in Pro(35S):ETR1-1 lines and corolla abscission did not take place. When senescence of Pro(35S):ETR1-1 plants did take place this was accompanied by leaf bleaching, but tissues remained fully turgid and showed no signs of collapse. A single Pro(35S):LeEIL1 line was found to exhibit consistently accelerated leaf and flower senescence and precocious flower bud shedding. CONCLUSIONS: These observations support a role for ethylene in regulating a spectrum of developmental events associated with organ senescence and tissue necrosis. Furthermore, the transgenic lines generated during this study may provide a valuable resource for exploring how senescence processes are regulated in plants.     

Soluble carbohydrates in Delphinium and their influence on sepal abscission in cut flowers

A carbohydrate other than sucrose, glucose, fructose and myo -inositol was detected in sepal extracts of Delphinium . This compound was identified as mannitol by ¹H-NMR. Mannitol was the major carbohydrate in all examined organs: the sepal, the other parts of the flower, the stem and leaves. Mannitol as well as glucose (both at 0.55 M ), fed to cut Delphinium flowers, similarly delayed the abscission of sepals. 3- O -methyl glucose (3-OMG) and polyethylene glycol 200 at the same molar concentrations had no such effect. The treatment with glucose markedly increased the concentrations of glucose and fructose in the sepals without changing the concentrations of sucrose and mannitol. On the other hand, the treatment with mannitol increased the concentrations of glucose and fructose in addition to mannitol in the sepals, suggesting that mannitol is metabolized in Delphinium flowers. The treatment with 3-OMG increased the concentration of 3-OMG but not other carbohydrates. Mannitol and glucose similarly delayed the increase in ethylene production in flowers, but 3-OMG did not. The sensitivity to ethylene was similarly reduced by the treatment with glucose and mannitol, but not by 3-OMG. These results suggest that the treatment with mannitol, a major carbohydrate in Delphinium , delayed the abscission of sepals by reducing the sensitivity to ethylene. Mannitol further acted, not merely as an osmolyte, but as an apparent source for carbohydrate metabolism in the flower.     

The never ripe mutation blocks ethylene perception in tomato.

Seedlings of tomato fruit ripening mutants were screened for their ability to respond to ethylene. Ethylene induced the triple response in etiolated hypocotyls of all tomato ripening mutants tested except for one, Never ripe (Nr). Our results indicated that the lack of ripening in this mutant is caused by ethylene insensitivity. Segregation analysis indicated that Nr-associated ethylene insensitivity is a single codominant trait and is pleiotropic, blocking senescence and abscission of flowers and the epinastic response of petioles. In normal tomato flowers, petal abscission and senescence occur 4 to 5 days after the flower opens and precede fruit expansion. If fertilization does not occur, pedicel abscission occurs 5 to 8 days after petal senescence. If unfertilized, Nr flowers remained attached to the plant indefinitely, and petals remained viable and turgid more than four times longer than their normal counterparts. Fruit development in Nr plants was not preceded by petal senescence; petals and anthers remained attached until they were physically displaced by the expanding ovary. Analysis of engineered 1-aminocyclopropane-1-carboxylate (ACC) synthase-overexpressing plants indicated that they are phenotypic opposites of Nr plants. Constitutive expression of ACC synthase in tomato plants resulted in high rates of ethylene production by many tissues of the plant and induced petiole epinasty and premature senescence and abscission of flowers, usually before anthesis. There were no obvious effects on senescence in leaves of ACC synthase overexpressers, suggesting that although ethylene may be important, it is not sufficient to cause tomato leaf senescence; other signals are clearly involved.     

Further examination of abscission zone cells as ethylene target cells in higher plants

BACKGROUND AND AIMS: Two aspects of the competence of abscission zone cells as a specific class of hormone target cell are examined. The first is the competence of these target cells to respond to a remote stele-generated signal, and whether ethylene acts in concert with this signal to initiate abscission of the primary leaf in Phaseolus vulgaris. The second is to extend the concept of dual control of abscission cell competence. Can the concept of developmental memory that is retained by abscission cell of Phaseolus vulgaris post-separation in terms of the inductive/repressive control of beta-1,4-glucan endohydrolase (cellulase) activity exerted by ethylene/auxin be extended to the rachis abscission zone cells of Sambucus nigra? METHODS: Abscission assays were performed using the leaf petiole-pulvinus explants of P. vulgaris with the distal pulvinus stele removed. These (-stele) explants do not separate when treated with ethylene and require a stele-generated signal from the distal pulvinus for separation at the leaf petiole-pulvinis abscission zone. Using these explants, the role of ethylene was examined, using the ethylene action blocker, 1-methyl cyclopropene, as well as the significance of the tissue from which the stele signal originates. Further, leaf rachis abscission explants were excised from the compound leaves of S. nigra, and changes in the activity of cellulase in response to added ethylene and auxin post-separation was examined. KEY RESULTS: The use of (-stele) explants has confirmed that ethylene, with the stele-generated signal, is essential for abscission. Neither ethylene alone nor the stelar signal alone is sufficient. Further, in addition to the leaf pulvinus distal to the abscission zone, mid-rib tissue that is excised from senescent or green mid-rib tissue can also generate a competent stelar signal. Experiments with rachis abscission explants of S. nigra have shown that auxin, when added to cells post-separation can retard cellulase activity, with activity re-established with subsequent ethylene treatment. CONCLUSIONS: The triggers that initiate and regulate the separation process are complex with, in bean leaves at least, the generation of a signal (or signals) from remote tissues, in concert with ethylene, a requisite part of the process. Once evoked, abscission cells maintain a developmental memory such that the induction/repression mediated by ethylene/auxin that is observed prior to separation is also retained by the cells post-separation.     

Use of Mutants to Dissect the Role of Ethylene Signalling in Organ Senescence and the Regulation of Yield in Arabidopsis thaliana

The role of ethylene in regulating organ senescence in Arabidopsis has been investigated by studying the development of mutants that have an attenuated capacity to perceive the gas. The onset of leaf senescence and floral organ abscission was delayed in the ethylene-insensitive mutant etr1. The photosynthetic life span of rosette leaves was similarly extended in the gain-of-function mutant ers2, and this mutant also exhibited a delay in the timing of pod dehiscence primarily as a consequence of an extension in the final stages of senescence. A detailed analysis of yield revealed that whilst thousand grain weight was increased, by as much as 20 %, in etr1, ein4, and the loss-of-function mutant etr2, only the latter showed a significant increase in total weight of seeds produced per plant. The other studied mutants exhibited a reduction in total seed yield of almost 40 %. These observations are discussed in the context of the possible role of ethylene in regulating organ senescence and their significance in the breeding of crop plants with enhanced phenotypic characteristics.