Cap Weakening Research Articles

Tissue-specific pectin methylesterification and pectin methylesterase activities play a role in lettuce seed germination

Cell wall mechanical strength is partially controlled by the degree of pectin methylesterification that is mediated through the action of pectin methylesterases (PMEs). Changes in cell wall mechanical strength are involved in the endosperm cap weakening and radicle elongation growth during the germination of some endospermic seeds. However, little is known about the roles of the status of pectin methylesterification and PME activities during seed germination, especially in lettuce, a traditional model plant for seed germination research. In the present study, we demonstrated that the rupture of lettuce endosperm cap during seed germination was caused by cell rupture, not by cell separation as formerly reported in other species. Next, we investigated the status of pectin methylesterification of the endosperm cap and radicle during germination, using immunolabeling by specific monoclonal antibodies against methyl-esterified pectins and calcium bridges. We also analyzed the expression patterns of PME activities and an encoding gene LsPME/PMEI51 in the endosperm cap and radicle. It was found that cell walls of the endosperm cap comprise more methyl-esterified pectins than radicle, while PME activities and the abundance of LsPME/PMEI51 transcripts in the endosperm cap are lower than those in the radicle. In addition, the status of pectin methylesterification and PME expression patterns changed during seed germination. Low PME activities and high pectin methylesterification in the endosperm cap is hypothesized to be the reason of why endosperm does not rupture by cell separation. It is concluded that tissue-specific pectin methylesterification and PME activities play a role in lettuce seed germination.

Nicotine Upregulates Metalloproteinase Function in Vascular Smooth Muscle Cells Inducing Cell Migration

Abstract Objectives We hypothesize that nicotine plays a role in the progression of atherosclerosis by inducing VSMC phenotypic switching and cell migration from the media to an inner layer–the intima. This forms a new layer called the neointima, which becomes a plaque with a lipid-rich necrotic core and fibrous VSMC cap. The cap confers protection against plaque rupture and stroke or heart attack. However, VSMCs can contribute to cap thinning by secreting metalloproteinases (MMPs) which degrade the extracellular matrix (ECM) creating a plaque prone to rupture. Methods Our lab will test blackberry extract. Blackberries contain health-promoting phytochemicals called polyphenols and other polyphenol extracts have been shown to inhibit MMPs. Specifically, blackberry extract will be added to nicotine treated cells and to animal model diets. Results In VSMCs, we found that nicotine significantly increased the expression of several MMPs, while decreasing the expression of multiple tissue inhibitors of metalloproteinases (TIMPs). MMP activity measured by zymography was also upregulated by nicotine suggesting that nicotine modulates extracellular matrix (ECM) remodeling by increasing MMP expression and activity. Consistent with ECM remodeling, nicotine promoted cell migration, which was associated with increased reactive oxygen species (ROS) levels. Conclusions These data suggest nicotine promotes VSMC migration into the intima and contributes to plaque instability by fibrous cap weakening. However, future studies are needed to demonstrate nicotine changes plaque composition in vivo. To negate detrimental effects, our lab will test blackberry extract. Blackberries contain health-promoting phytochemicals called polyphenols and other polyphenol extracts have been shown to inhibit MMPs. Specifically, blackberry extract will be added to nicotine treated cells and to animal model diets. Funding Sources Florida Department of Health.

Abscisic acid and ethephon regulation of cellulase in the endosperm cap and radicle during lettuce seed germination.

The purpose of this study was to investigate the role of cellulase in endosperm cap weakening and radicle elongation during lettuce (Lactuca sativa L.) seed germination. The application of abscisic acid (ABA) or ethephon inhibits or promotes germination, respectively, by affecting endosperm cap weakening and radicle elongation. Cellulase activities, and related protein and transcript abundances of two lettuce cellulase genes, LsCEL1 and LsCEL2, increase in the endosperm cap and radicle prior to radicle protrusion following imbibition in water. ABA or ethephon reduce or elevate, respectively, cellulase activity, and related protein and transcript abundances in the endosperm cap. Taken together, these observations suggest that cellulase plays a role in endosperm cap weakening and radicle elongation during lettuce seed germination, and that the regulation of cellulase in the endosperm cap by ABA and ethephon play a role in endosperm cap weakening. However, the influence of ABA and ethephon on radicle elongation may not be through their effects on cellulase.

Suppression of α-l-arabinofuranosidase in the endosperm and atypical germination of lettuce seeds induced by sodium dichloroisocyanurate

Endosperm cap (CAP) weakening and radicle (RAD) elongation growth are prerequisites for the completion of lettuce seed germination. Lettuce CAP and RAD cell walls contain a large amount of l-arabinofuranose, which may exist in the form of arabinan or l-arabinofuranose-rich polysaccharides. The aim of the present study was to investigate the relationship between α-l-arabinofuranosidase, an enzyme degrading l-arabinofuranose-rich polysaccharides, and CAP weakening and RAD elongation growth. It was shown that the enzyme activity and the transcript abundance of two lettuce α-l-arabinofuranosidase genes, LsARAf2 and LsARAf7, increased in the CAP and RAD prior to the completion of seed germination during imbibition in water. Sodium dichloroisocyanurate, a chemical inducing atypical germination (RAD elongation without endosperm rupture) in lettuce seeds, significantly decreased the enzyme activity and the transcript abundance of LsARAf7 and LsARAf2 in the CAP. It is concluded that the expression of α-l-arabinofuranosidase is necessary for CAP weakening and the suppression of this enzyme in the CAP is at least partly responsible for atypical germination in lettuce.

Involvement of reactive oxygen species in endosperm cap weakening and embryo elongation growth during lettuce seed germination.

Endosperm cap (CAP) weakening and embryo elongation growth are prerequisites for the completion of lettuce seed germination. Although it has been proposed that the cell wall loosening underlying these processes results from an enzymatic mechanism, it is still unclear which enzymes are involved. Here it is shown that reactive oxygen species (ROS), which are non-enzymatic factors, may be involved in the two processes. In Guasihong lettuce seeds imbibed in water, O2·(-) and H2O2 accumulated and peroxidase activity increased in the CAP, whereas its puncture force decreased. In addition, in the radicle, the increase in embryo growth potential was accompanied by accumulation of O2·(-) and an increase in peroxidase activity. Imbibing seeds in 0.3% sodium dichloroisocyanurate (SDIC) reduced endosperm viability and the levels of O2·(-), H2O2, and peroxidase activity in the CAP, whereas the decrease in its puncture force was inhibited. However, in the embryo, SDIC did not affect the accumulation of O2·(-), peroxidase activity, and the embryo growth potential. As a result, SDIC caused atypical germination, in which the endosperm ruptured at the boundary between the CAP and lateral endosperm. ROS scavengers and ROS generation inhibitors inhibited the CAP weakening and also decreased the embryo growth potential, thus decreasing the percentage of seed germination. Exogenous ROS and ROS generation inducers increased the percentage of CAP rupture to some extent, and the addition of H2O2 to 0.3% SDIC enabled some seeds to undergo typical germination.

ABA crosstalk with ethylene and nitric oxide in seed dormancy and germination.

Dormancy is an adaptive trait that enables seed germination to coincide with favorable environmental conditions. It has been clearly demonstrated that dormancy is induced by abscisic acid (ABA) during seed development on the mother plant. After seed dispersal, germination is preceded by a decline in ABA in imbibed seeds, which results from ABA catabolism through 8′-hydroxylation. The hormonal balance between ABA and gibberellins (GAs) has been shown to act as an integrator of environmental cues to maintain dormancy or activate germination. The interplay of ABA with other endogenous signals is however less documented. In numerous species, ethylene counteracts ABA signaling pathways and induces germination. In Brassicaceae seeds, ethylene prevents the inhibitory effects of ABA on endosperm cap weakening, thereby facilitating endosperm rupture and radicle emergence. Moreover, enhanced seed dormancy in Arabidopsis ethylene-insensitive mutants results from greater ABA sensitivity. Conversely, ABA limits ethylene action by down-regulating its biosynthesis. Nitric oxide (NO) has been proposed as a common actor in the ABA and ethylene crosstalk in seed. Indeed, convergent evidence indicates that NO is produced rapidly after seed imbibition and promotes germination by inducing the expression of the ABA 8′-hydroxylase gene, CYP707A2, and stimulating ethylene production. The role of NO and other nitrogen-containing compounds, such as nitrate, in seed dormancy breakage and germination stimulation has been reported in several species. This review will describe our current knowledge of ABA crosstalk with ethylene and NO, both volatile compounds that have been shown to counteract ABA action in seeds and to improve dormancy release and germination.

Molecular and physiological dissection of enhanced seed germination using short-term low-concentration salt seed priming in tomato

Seed germination is the initial step of plant development. Seed priming with salt promotes seed germination in tomato (Solanum lycopersicum L.); however, the molecular and physiological mechanisms underlying the enhancement of seed germination by priming remain to be elucidated. In this study, we examined the following in seeds both during and after priming treatment: the endogenous abscisic acid (ABA) and gibberellin (GA) concentrations; the expression of genes encoding ABA catabolic and GA biosynthesis enzymes, including 8'-hydroxylase (CYP707A), copalyl diphosphate synthase (CPS), GA 20-oxidase (GA20ox) and GA 3-oxidase (GA3ox); and endosperm cap weakening enzymes, including expansin (EXP), class I β-1,3-glucanase (GulB), endo-β-mannanase (MAN) and xyloglucan endotransglucosylase (XTH). Tomato seeds were soaked for 24h at 25°C in the dark in 300mM NaCl (NaCl-priming) or distilled water (hydro-priming). For both priming treatments, the ABA content in the seeds increased during treatment but rapidly decreased after sowing. Both during and after the priming treatments, the ABA levels in the hydro-primed seeds and NaCl-primed seeds were not significantly different. The expression levels of SlGA20ox1, SlGA3ox1 and SlGA3ox2 were significantly enhanced in the NaCl-primed seeds compared to the hydro-primed seeds. The GA(4) content was quantifiable after both types of priming, indicating that GA(4) is the major bioactive GA molecule involved in tomato seed germination. The GA(4) content was significantly higher in the NaCl-primed seeds than in the hydro-primed seeds 12h after sowing and thereafter. Additionally, the peak expression levels of SlEXP4, SlGulB, SlMAN2 and SlXTH4 occurred earlier and were significantly higher in the NaCl-primed seeds than in the hydro-primed seeds. These results suggest that the observed effect of NaCl-priming on tomato seed germination is caused by an increase of the GA(4) content via GA biosynthetic gene activation and a subsequent increase in the expression of genes related to endosperm cap weakening.

Regulation of seed germination in the close Arabidopsis relative Lepidium sativum: a global tissue-specific transcript analysis.

The completion of germination in Lepidium sativum and other endospermic seeds (e.g. Arabidopsis [Arabidopsis thaliana]) is regulated by two opposing forces, the growth potential of the radicle (RAD) and the resistance to this growth from the micropylar endosperm cap (CAP) surrounding it. We show by puncture force measurement that the CAP progressively weakens during germination, and we have conducted a time-course transcript analysis of RAD and CAP tissues throughout this process. We have also used specific inhibitors to investigate the importance of transcription, translation, and posttranslation levels of regulation of endosperm weakening in isolated CAPs. Although the impact of inhibiting translation is greater, both transcription and translation are required for the completion of endosperm weakening in the whole seed population. The majority of genes expressed during this process occur in both tissues, but where they are uniquely expressed, or significantly differentially expressed between tissues, this relates to the functions of the RAD as growing tissue and the CAP as a regulator of germination through weakening. More detailed analysis showed that putative orthologs of cell wall-remodeling genes are expressed in a complex manner during CAP weakening, suggesting distinct roles in the RAD and CAP. Expression patterns are also consistent with the CAP being a receptor for environmental signals influencing germination. Inhibitors of the aspartic, serine, and cysteine proteases reduced the number of isolated CAPs in which weakening developed, and inhibition of the 26S proteasome resulted in its complete cessation. This indicates that targeted protein degradation is a major control point for endosperm weakening.

Ethylene Interacts with Abscisic Acid to Regulate Endosperm Rupture during Germination: A Comparative Approach UsingLepidium sativumandArabidopsis thaliana

The micropylar endosperm cap covering the radicle in the mature seeds of most angiosperms acts as a constraint that regulates seed germination. Here, we report on a comparative seed biology study with the close Brassicaceae relatives Lepidium sativum and Arabidopsis thaliana showing that ethylene biosynthesis and signaling regulate seed germination by a mechanism that requires the coordinated action of the radicle and the endosperm cap. The larger seed size of Lepidium allows direct tissue-specific biomechanical, biochemical, and transcriptome analyses. We show that ethylene promotes endosperm cap weakening of Lepidium and endosperm rupture of both species and that it counteracts the inhibitory action of abscisic acid (ABA) on these two processes. Cross-species microarrays of the Lepidium micropylar endosperm cap and the radicle show that the ethylene-ABA antagonism involves both tissues and has the micropylar endosperm cap as a major target. Ethylene counteracts the ABA-induced inhibition without affecting seed ABA levels. The Arabidopsis loss-of-function mutants ACC oxidase2 (aco2; ethylene biosynthesis) and constitutive triple response1 (ethylene signaling) are impaired in the 1-aminocyclopropane-1-carboxylic acid (ACC)-mediated reversion of the ABA-induced inhibition of seed germination. Ethylene production by the ACC oxidase orthologs Lepidium ACO2 and Arabidopsis ACO2 appears to be a key regulatory step. Endosperm cap weakening and rupture are promoted by ethylene and inhibited by ABA to regulate germination in a process conserved across the Brassicaceae.

Statins inhibit cyclooxygenase-2 and matrix metalloproteinase-9 in human endothelial cells: anti-angiogenic actions possibly contributing to plaque stability

Cyclooxygenase (COX)-2 expression is increased in inflammation and angiogenesis and also in atherosclerotic plaques, where it co-localizes with metalloproteinases (MMPs) involved in the fibrous cap weakening. Insight into the regulation of COX-2 and MMP-9 expression suggests the involvement of a Rho-dependent pathway. Because statins interfere with Rho activation, we investigated the statin effect on COX-2 and MMP expressions in the human endothelium. Simvastatin and atorvastatin were incubated with endothelial cells for 12 h before stimulation with phorbol myristate acetate or tumour necrosis factor-alpha, for times suitable to assess the endothelial tube differentiation on Matrigel and COX-2 and MMPs activities, proteins, and mRNA expressions. At 0.1-10 micromol/L, both statins reduced COX-2 expression and activity, without affecting COX-1. The statin effect was reversed by mevalonate and geranylgeranyl-pyrophosphate and mimicked by the Rho inhibitor C3 transferase, indicating the involvement of Rho in the signal transduction pathway leading to COX-2 expression. In parallel, statins, as well as COX-2 inhibitors, reduced the MMP-9 stimulated release and the endothelial tubular differentiation. In the human vascular endothelium, statins reduce COX-2 and MMP-9 expression and activity. Through this mechanism, statins exert an anti-angiogenic effect possibly contributing to the cholesterol-lowering-unrelated protective effects of statins against plaque inflammatory angiogenesis and rupture.

Transcriptional profiling of cell wall protein genes in chickpea embryonic axes during germination and growth

Cell wall hydrolases have been assumed to be involved in the regulation of seed germination, mostly through their contribution to the cell wall disassembly associated with endosperm cap weakening. In Cicer arietinum (a non-endospermic leguminosae seed), we have focused our research directly on the elongation process of the embryonic axes themselves during germination. The genes encoding cell wall proteins, previously implicated in the elongation of chickpea epicotyls, might also be involved in the expansion of embryonic axis cells, and the modulation of their expression could be part of the control of the germinative process. Thus, chickpea α-expansins and xyloglucan endotransglycosylase/hydrolase (XTH) acting on the cellulose/xyloglucan network seem to be involved in the elongation of both chickpea epicotyls and embryonic axes, although the products of different genes perform their actions on each organ. Among the four known cDNAs encoding chickpea α-expansins, Ca-EXPA1 was the only isoform highly expressed in embryonic axes during germination. In contrast to epicotyl elongation, the genes encoding cell wall β-galactosidases, involved in pectin degradation, were not expressed during germination, suggesting no role in embryonic axis elongation, mainly due to the different metabolism of pectins during cell wall loosening in embryonic axis or epicotyl cells. The results concerning CanST-1 and -2, encoding two growth-related cell wall proteins, suggested that these genes were not involved in elongation of embryonic axes during germination. The transcription pattern of Cap28, which encodes a glutamic acid rich cell wall protein of unknown function, indicated a role in the development of the embryonic axes during germination.

Pregnancy-Associated Plasma Protein-A: Do Specific Markers of Vascular or Plaque Activation Exist, and Do We Really Need Them?

Acute coronary syndromes (ACS), which include unstable angina, myocardial infarction, and sudden cardiac death, continue to be a significant public health problem in industrialized countries. A growing body of research is directed at understanding the etiology of ACS and at identifying biomarkers of high risk. The results from angiographic and angioscopic studies suggest that the disruption of an atherosclerotic plaque with subsequent thrombosis may represent a common, although not the only, pathogenetic mechanism for ACS (1). The definitive reasons for plaque rupture are unknown, but inflammation appears to play a major role (2). Indeed, the main features that distinguish unstable from stable plaques include the presence of wide inflammatory infiltration, plaque fissures, and fresh thrombi (3). Early studies reported evidence of local activation of inflammatory cells in the shoulder region of coronary plaques, with release of proteolytic enzymes (metalloproteinases) that degrade the extracellular matrix and contribute to the fibrous cap weakening and plaque instability (1)(2)(3). More recently, however, it has been demonstrated that patients with ACS have widespread inflammation of the coronary tree as well as systemic activation of the inflammatory system (4)(5). These findings have stimulated the search for an ideal marker of plaque instability. Liuzzo et al. (5) reported for the first time that systemic concentrations of C-reactive protein (CRP), a prototypic marker of inflammation, were increased in a population of patients with unstable angina and were associated with a high recurrence of cardiac events. Larger studies confirmed the predictive value of …

Exogenous gibberellins inhibit coffee (Coffea arabica cv. Rubi) seed germination and cause cell death in the embryo

The mechanism of inhibition of coffee (Coffea arabica cv. Rubi) seed germination by exogenous gibberellins (GAs) and the requirement of germination for endogenous GA were studied. Exogenous GA(4+7) inhibited coffee seed germination. The response to GA(4+7) showed two sensitivity thresholds: a lower one between 0 and 1 microM and a higher one between 10 and 100 microM. However, radicle protrusion in coffee seed depended on the de novo synthesis of GAs. Endogenous GAs were required for embryo cell elongation and endosperm cap weakening. Incubation of coffee seed in exogenous GA(4+7) led to loss of embryo viability and dead cells were observed by low temperature scanning microscopy only when the endosperm was surrounding the embryo. The results described here indicate that the inhibition of germination by exogenous GAs is caused by factors that are released from the endosperm during or after its weakening, causing cell death in the embryo and leading to inhibition of radicle protrusion.

Abscisic acid controls embryo growth potential and endosperm cap weakening during coffee (Coffea arabica cv. Rubi) seed germination.

The mechanism and regulation of coffee seed germination were studied in Coffea arabica L. cv. Rubi. The coffee embryo grew inside the endosperm prior to radicle protrusion and abscisic acid (ABA) inhibited the increase in its pressure potential. There were two steps of endosperm cap weakening. An increase in cellulase activity coincided with the first step and an increase in endo-beta-mannanase (EBM) activity with the second step. ABA inhibited the second step of endosperm cap weakening, presumably by inhibiting the activities of at least two EBM isoforms and/or, indirectly, by inhibiting the pressure force of the radicle. The increase in the activities of EBM and cellulase coincided with the decrease in the force required to puncture the endosperm and with the appearance of porosity in the cell walls as observed by low-temperature scanning electronic microscopy. Tissue printing showed that EBM activity was spatially regulated in the endosperm. Activity was initiated in the endosperm cap whereas later during germination it could also be detected in the remainder of the endosperm. Tissue printing revealed that ABA inhibited most of the EBM activity in the endosperm cap, but not in the remainder of the endosperm. ABA did not inhibit cellulase activity. There was a transient rise in ABA content in the embryo during imbibition, which was likely to be responsible for slow germination, suggesting that endogenous ABA also may control embryo growth potential and the second step of endosperm cap weakening during coffee seed germination.

Class I chitinase and beta-1,3-glucanase are differentially regulated by wounding, methyl jasmonate, ethylene, and gibberellin in tomato seeds and leaves.

Class I chitinase (Chi9) and beta-1,3-glucanase (GluB) genes are expressed in the micropylar endosperm cap of tomato (Lycopersicon esculentum) seeds just before radicle emergence through this tissue to complete germination. In gibberellin (GA)-deficient mutant (gib-1) seeds, expression of Chi9 and GluB mRNA and protein is dependent upon GA. However, as expression occurs relatively late in the germination process, we investigated whether the genes are induced indirectly in response to tissue wounding associated with endosperm cap weakening and radicle protrusion. Wounding and methyl jasmonate (MeJA) induced Chi9 expression, whereas ethylene, abscisic acid, sodium salicylate, fusicoccin, or beta-aminobutyric acid were without effect. Chi9 expression occurred only in the micropylar tissues when seeds were exposed to MeJA or were wounded at the chalazal end of the seed. Expression of Chi9, but not GluB, mRNA was reduced in germinating seeds of the jasmonate-deficient defenseless1 tomato mutant and could be restored by MeJA treatment. Chi9 expression during germination may be associated with "wounding" from cell wall hydrolysis and weakening in the endosperm cap leading to radicle protrusion, and jasmonate is involved in the signaling pathway for this response. Among these treatments and chemicals (other than GA), only MeJA and wounding induced a low level of GluB expression in gib-1 seeds. However, MeJA, wounding, and particularly ethylene induced both genes in leaves, whereas GA induced only Chi9 in leaves. Although normally expressed simultaneously during tomato seed germination, Chi9 and GluB genes are regulated distinctly and tissue specifically by hormones and wounding.

A gibberellin-regulated xyloglucan endotransglycosylase gene is expressed in the endosperm cap during tomato seed germination.

Xyloglucan endotransglycosylases (XETs) modify xyloglucans, major components of primary cell walls in dicots. A cDNA encoding an XET (LeXET4) was isolated from a germinating tomato (Lycopersicon esculentum Mill.) seed cDNA library. DNA gel blot analysis showed that LeXET4 is a single-copy gene in the tomato genome. LeXET4 mRNA was strongly expressed in germinating seeds, was much less abundant in stems, and was not detected in roots, leaves or flower tissues. During germination, LeXET4 mRNA was detected in seeds within 12 h of imbibition with maximum mRNA abundance at 24 h. Tissue prints showed that LeXET4 mRNA was localized exclusively to the endosperm cap region. Expression of LeXET4 was dependent on exogenous gibberellin (GA) in GA-deficient (gib-1 mutant) tomato seeds, while abscisic acid, a seed germination inhibitor, had no effect on LeXET4 mRNA expression in wild-type seeds. LeXET4 mRNA disappeared after radicle emergence, even though degradation of the lateral endosperm cell walls continued. The temporal, spatial and hormonal regulation pattern of LeXET4 gene expression suggests that XET has a role in endosperm cap weakening, a key process regulating tomato seed germination.

Class I beta-1,3-glucanase and chitinase are expressed in the micropylar endosperm of tomato seeds prior to radicle emergence.

beta-1,3-Glucanase (EC 3.2.1.39) and chitinase (EC 3.2.1.14) mRNAs, proteins, and enzyme activities were expressed specifically in the micropylar tissues of imbibed tomato (Lycopersicon esculentum Mill.) seeds prior to radicle emergence. RNA hybridization and immunoblotting demonstrated that both enzymes were class I basic isoforms. beta-1,3-Glucanase was expressed exclusively in the endosperm cap tissue, whereas chitinase localized to both endosperm cap and radicle tip tissues. beta-1,3-Glucanase and chitinase appeared in the micropylar tissues of gibberellin-deficient gib-1 tomato seeds only when supplied with gibberellin. Accumulation of beta-1,3-glucanase mRNA, protein and enzyme activity was reduced by 100 microM abscisic acid, which delayed or prevented radicle emergence but not endosperm cap weakening. In contrast, expression of chitinase mRNA, protein, and enzyme activity was not affected by abscisic acid. Neither of these enzymes significantly hydrolyzed isolated tomato endosperm cap cell walls. Although both beta-1,3-glucanase and chitinase were expressed in tomato endosperm cap tissue prior to radicle emergence, we found no evidence that they were directly involved in cell wall modification or tissue weakening. Possible functions of these hydrolases during tomato seed germination are discussed.

Expression of an expansin is associated with endosperm weakening during tomato seed germination.

Expansins are extracellular proteins that facilitate cell wall extension, possibly by disrupting hydrogen bonding between hemicellulosic wall components and cellulose microfibrils. In addition, some expansins are expressed in non-growing tissues such as ripening fruits, where they may contribute to cell wall disassembly associated with tissue softening. We have identified at least three expansin genes that are expressed in tomato (Lycopersicon esculentum Mill.) seeds during germination. Among these, LeEXP4 mRNA is specifically localized to the micropylar endosperm cap region, suggesting that the protein might contribute to tissue weakening that is required for radicle emergence. In gibberellin (GA)-deficient (gib-1) mutant seeds, which germinate only in the presence of exogenous GA, GA induces the expression of LeEXP4 within 12 hours of imbibition. When gib-1 seeds were imbibed in GA solution combined with 100 microM abscisic acid, the expression of LeEXP4 was not reduced, although radicle emergence was inhibited. In wild-type seeds, LeEXP4 mRNA accumulation was blocked by far-red light and decreased by low water potential but was not affected by abscisic acid. The presence of LeEXP4 mRNA during seed germination parallels endosperm cap weakening determined by puncture force analysis. We hypothesize that LeEXP4 is involved in the regulation of seed germination by contributing to cell wall disassembly associated with endosperm cap weakening.

The second step of the biphasic endosperm cap weakening that mediates tomato ( Lycopersicon esculentum ) seed germination is under control of ABA

The role of abscisic acid (ABA) in the weakening of the endosperm cap prior to radicle protrusion in tomato ( Lycopersicon esculentum Mill. cv. Moneymaker) seeds was studied. The endosperm cap weakened substantially in both water and ABA during the first 38 h of imbibition. After 38 h the force required for endosperm cap puncturing was arrested at 0.35 N in ABA, whereas in water a further decrease occurred until the radicle protruded. During the first 2 d of imbibition endo‐β‐mannanase activity was correlated with the decrease in required puncture force and with the appearance of ice‐crystal‐induced porosity in the cell walls as observed by scanning electron microscopy. Prolonged incubation in ABA resulted in the loss of endo‐β‐mannanase activity and the loss of ice‐crystal‐induced porosity, but not in a reversion of the required puncture force. ABA also had a distinct but minor effect on the growth potential of the embryo. However, endosperm cap resistance played the limiting role in the completion of germination. It was concluded that (a) endosperm cap weakening is a biphasic process and (b) inhibition of germination by ABA is through the second step in the endosperm cap weakening process.

The second step of the biphasic endosperm cap weakening that mediates tomato (Lycopersicon esculentum) seed germination is under control of ABA

The role of abscisic acid (ABA) in the weakening of the endosperm cap prior to radicle protrusion in tomato (Lycopersicon esculentum Mill. cv. Moneymaker) seeds was studied. The endosperm cap weakened substantially in both water and ABA during the first 38 h of imbibition. After 38 h the force required for endosperm cap puncturing was arrested at 0.35 N in ABA, whereas in water a further decrease occurred until the radicle protruded. During the first 2 d of imbibition endo-beta-mannanase activity was correlated with the decrease in required puncture force and with the appearance of ice-crystal-induced porosity in the cell walls as observed by scanning electron microscopy. Prolonged incubation in ABA resulted in the loss of endo-beta-mannanase activity and the loss of ice-crystal-induced porosity, but not in a reversion of the required puncture force. ABA also had a distinct but minor effect on the growth potential of the embryo. However, endosperm cap resistance played the limiting role in the completion of germination. It was concluded that (a) endosperm cap weakening is a biphasic process and (b) inhibition of germination by ABA is through the second step in the endosperm cap weakening process.