Journal of Experimental Botany, Vol. 53, No. 374, pp. 1643-1650,
July 1, 2002
© 2002 Oxford University Press
Peroxidase activity develops in the micropylar endosperm of tomato seeds prior to radicle protrusion
Received 2 November 2001; Accepted 22 March 2002
1 Department of Regulatory Biology, Faculty of Science, Saitama University, Urawa, Saitama 338-8570, Japan
1 Fax: +81 48 858 3584. E-mail: moro{at}post.saitama-u.ac.jp
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Peroxidase activity developed specifically in the micropylar region of the endosperm of imbibed tomato seeds prior to radicle emergence. The activity was first detected approximately 24 h after the start of imibibition (6 h before radicle emergence) and increased markedly thereafter. In the lateral portion of the endosperm, peroxidase activity was undetectable for the first 2 d after the start of imbibition. Although the activity in the lateral endosperm became detectable 3 d after imbibition, the extent of the development of the activity was slight. The localization of peroxidase activity in the micropylar endosperm 2 d after the start of imbibition was confirmed by tissue printing analyses. When the endosperm tissues were wounded, there was an enhancement of the enzyme activity at the wounded region. H2O2 was formed at the expense of NADH only in the presence of Mn2+ and dinitrophenol by the extract from the micropylar endosperm in which peroxidase activity was present. The presence of H2O2 in the micropylar portion of the endosperm was shown histochemically. The possible functions of the peroxidases that develop in the endosperm of tomato seeds are discussed.
Key words: Key words: Endosperm, germination, hydrogen peroxide, peroxidase, tomato seed.
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
It was reported in a previous paper that the activity of a basic ß-1,3-glucanase develops markedly in the micropylar region of the endosperm of germinated tomato seeds (Morohashi and Matsushima, 2000). Wu et al. (2001) have recently shown by using a more sensitive assay method that enzyme activity is already expressed prior to radicle emergence. However, the physiological significance of the development of the enzyme in the micropylar tissues of germinating and germinated tomato seeds is not well understood at present. In tobacco seeds, Vogeli-Lange et al. (1994) and Leubner-Metzger et al. (1995) have shown that ß-1,3-glucanase is induced in the micropylar part of the endosperm prior to radicle protrusion and have suggested that the enzyme contributes to the weakening of the cell walls of the micropylar endosperm by helping to hydrolyse cell wall ß-1,3-glucans and thereby contributes to facilitating penetration of the radicle. However, substrates for ß-1,3-glucanase do not seem to be present in the cell wall of the endosperm of tomato seeds (Wu et al., 2001). Therefore, it is unlikely with tomato seeds that the enzyme contributes to the weakening of the cell walls of the micropylar part of the endosperm.
When the radicle of tomato seeds penetrates the micropylar portion of the endosperm, that endosperm part is ruptured. The ruptured part will be an avenue for the invasion of pathogens. It is possible that some mechanism(s) is present for protecting the ruptured micropylar endosperm against pathogen entry. It is well known that ß-1,3-glucanase is induced as part of the defence reaction of plants to pathogen attack (Simmons, 1994). It has been reported that ß-1,3-glucanase is induced in response to wounding in plant tissues including tomato endosperm (Mauch et al., 1988; Derckel et al., 1998; Morohashi and Matsushima, 2000). Radicle penetration through the micropylar portion of the endosperm may accompany the breakdown of the cell wall, that is, wounding. The possibility, therefore, is suggested that ß-1,3-glucanase accumulating in the micropylar endosperm may play a role in protecting tomato seeds from pathogen infection through ruptured endosperm tissues, as has been proposed previously (Petruzzelli et al., 1999; Morohashi and Matsushima, 2000; Wu et al., 2001). It is known that chitinases are induced coordinately with ß-1,3-glucanases and act in synergy with ß-1,3-glucanases in protecting plants from pathogen infection (Simmons, 1994). Interestingly, Wu et al. (2001) have shown that chitinase is expressed, together with ß-1,3-glucanase, in the micropylar tissues of tomato seeds.
Many studies have shown that peroxidase is induced by wounding in plants (Desbiez and Boyer, 1981; Espelie et al., 1986; Svelheim and Robertsen, 1990; Kawaoka et al., 1994; Hiraga et al., 2000; Kato et al., 2000). Peroxidase is probably implicated in lignin and/or suberin formation during the polymerization of monoglinols or aromatic monomers (Kolattukudy, 1981; Whetten et al., 1998). Lignification and suberization was believed to play a role in the defensive responses to the entry of pathogenic microorganisms through a wounded part by developing physical barriers (Espelie et al., 1986; Kolattukudy et al., 1992; Siegel, 1993; Quiroga et al., 2000). On the other hand, peroxidase has been reported to be involved in generation of H2O2 from NADH (Maeder et al., 1980; Maeder and Amberg-Fisher, 1982). There are quite a few studies reporting that H2O2 serves as an important factor controlling pathogen-resistance responses and programmed cell death in plants (Lamb and Dixon, 1997; Wu et al., 1997; Bestwick et al., 1998; Joseph et al., 1998; Grant and Loake, 2000; DeRafael et al., 2001). Taking these into consideration, it is of interest to know if peroxidase, like ß-1,3-glucanase, is expressed specifically in the micropylar part of the endosperm of tomato seeds and whether it is induced by wounding in the endosperm. The pattern of the development of peroxidase activity in the endosperm of tomato seeds is reported here in connection with the possible physiological significance of the enzyme.
|
Materials and methods |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Plant material
Tomato (Lycopersicon esculentum [L.] Mill. cv. First Up) seeds, a kind gift from Sakata Seed Corp. (Yokohama, Japan), were used. For germination, they were placed on wet filter paper in Petri dishes and incubated at 25 °C in the dark.
Endosperm isolation
A seed was transversally cut into halves to produce the micropylar half-seed and the lateral half-seed. After cutting, the embryo parts were carefully removed by pushing out using forceps. The de-embryonated micropylar half-seed and lateral half-seed were denoted as micropylar endosperm half and lateral endosperm half, respectively, although they contained part of the testa. In some cases, the micropylar seed tip was excised from seeds as previously described (Nonogaki et al., 1992) and the embryo parts were pushed out with forceps. The de-embryonated micropylar seed tip was referred to as the micropylar endosperm. A wound was inflicted on the endosperm by cutting the tissue. One-day-imbibed seeds were transversally cut in the middle and the lateral half-seed was incubated on wet filter paper at 25 °C in the dark. After incubation for 1 d, embryo parts were removed from the wounded half-seed and the de-embyonated lateral half-seed (wounded lateral endosperm) was subjected to analyses for peroxidase activities and isoforms.
Enzyme extraction and assay
Endosperm parts (30 endosperm halves or 60 micropylar endosperms) were homogenized in 0.8–1.2 ml of 1/3x McIlvaine buffer (0.1 M citric acid–0.2 M disodium phosphate; pH 6) containing 1 M NaCl in a chilled mortar and pestle. The homogenate was centrifuged at 10 000 g for 2 min. The supernatant was used for enzyme assays. Peroxidase activity was assayed in 6 mM guaiacol, 4 mM H2O2 and 1/3x McIlvaine buffer (pH 6.8). The formation of tetraguaicol was followed by recording the absorbance at 470 nm. One unit of the enzyme activity represents the activity catalysing the formation of 0.01 µmol tetraguaiacol min–1. NADH oxidation was monitored by recording the absorbance at 340 nm in 50 mM Na-acetate buffer (pH 4.8), 0.15 mM NADH, 1 mM MnCl2 and 1 mM 2,4-dichlorophenol (DCP).
Isoelectric focusing
Endosperm extracts were dialysed against 1/10x McIlvaine buffer (pH 6) and were subjected to flat bed isoelectric focusing (IEF) on precast polyacrylamide gels in the pH range 3.5–9.5 (Ampholine PAGplate; Pharmacia Biotech) according to the manufacturer’s instructions. The peroxidase isozymes were visualized by soaking the gels in 20 mM Na-phosphate buffer (pH 6.8) containing 4-chloro-1-naphthol (0.6 mg ml–1) and 0.18% H2O2, according to Lagrimini and Rothstein (1987).
Tissue printing
Seeds imbibed for 1 d, seeds of 2-d-old seedlings and wounded lateral half-seeds were laterally cut into halves and the embryo parts were removed using forceps. De-embryonated seed parts were immediately laid with the cut surface down on the polyacrylamide gel (7.5%, w/v; 0.6 mm thick). After 5–10 min at room temperature (20–25 °C), tissues were removed from the gel and the gel was stained for peroxidase activity using 4-chloro-1-naphthol as the substrate, in the same way as in IEF gels.
Histochemical stain for monitoring the presence of H2O2
Laterally cut halves (de-embryonated) of seeds of 2-d-old seedlings were incubated for about 30 min at room temperature (20–25 °C) in a 1/10x McIlvaine buffer (pH 6) containing 1.2% (w/v) 3,5,3',5'-tetramethylbenzidine-HCl (TMB) (Barcelo, 1998). A blue staining appears at the sites where H2O2 accumulates.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Germination of tomato seeds
In the tomato seeds used in the present study, the radicle began to protrude approximately 30 h after the start of imbibition. Approximately 70% and 90% of seeds completed germination 40 h and 48 h after the start of imbibition, respectively.
Time-course of changes in peroxidase activity
Changes in peroxidase activities in the micropylar and the lateral endosperm half of tomato seeds were followed after the beginning of imbibition (Fig. 1). Peroxidase activity was below the limit of detection for up to 24 h in both the micropylar and the lateral endosperm half. After 1-d-imbibition, the activity, though very low, was detected in the micropylar endosperm half. The activity increased thereafter, especially after 30 h, and reached maximum at day 3. On the other hand, the activity in the lateral endosperm half remained at undetectable levels for the first 2 d. After that, it began to increase slightly.
|
To study in which part of the micropylar endosperm half the activity developed, the micropylar endosperm half of the seeds of 2-d-old seedlings was further divided transversally into three sections, micropylar end section (micropylar endosperm) and two consecutive sections approximately the same in width, and peroxidase activities in the micropylar endosperm and the section contiguous to the micropylar section were determined. As shown in Table 1A, high activity was detected in the micropylar endosperm of seeds of 2-d-old seedlings. By contrast, the activity in the contiguous section was below the limit of detection. In the endosperm of seeds of 3-d-old seedlings, on the other hand, peroxidase activity became detectable in the part contiguous to the micropylar endosperm (Table 1A). Taken together, it seems that peroxidase activity develops first in the micropylar endosperm and the active region spreads with time toward the lateral part. However, the majority of the activity in the endosperm was localized in the micropylar endosperm.
|
Effect of wounding on development of peroxidase activity
The effect of wounding on the development of peroxidase activity in the endosperm was studied by cutting the tissue (see Materials and methods). After 1 d of incubation, high activity of peroxidase was found in the wounded lateral endosperm (1.03±0.16 units part–1), while the activity was undetectable in the lateral endosperm half of intact seeds at the corresponding stage (2-d-old seedlings) (Fig. 1). This indicates that peroxidase activity is induced by wounding in the endosperm of tomato seeds. To study in which part of the wounded lateral endosperm the activity developed, the wounded lateral endosperm was transversally cut into halves to produce the section containing the wounded surface and the section away from the surface, and the activity in each section was determined. As seen from Table 1B. the activity was detected in the wounded region, but not in the part away from it. This indicates that the site where the peroxidase activity is induced by wounding is limited to the injured part and that the wounding effect is not transmitted to other parts away from the wounded region. This was confirmed by tissue printing analyses of the localization of peroxidase activity (see below).
NADH oxidation by the endosperm extract
The time-course of NADH oxidation by the extract from the micropylar endosperm of seeds of 2-d-old seedlings is shown in Fig. 2. The reaction was carried out in the presence of Mn2+ and DCP, because it has been shown that these cofactors enhance the peroxidase-catalysed oxidation of NADH (Halliwell, 1978). Requirement of these two factors for NADH oxidation by the endosperm extract was also confirmed in the present study; only slight oxidation of NADH occurred in the absence of either factor (data not shown). The oxidation of NADH was non-linear, and the rate was gradually accelerated with reaction time. This reaction pattern was similar to that reported with tobacco peroxidases (Maeder and Amberg-Fisher, 1982). H2O2 formation during NADH oxidation was monitored by adding guaiacol to the reaction mixture at the end of the reaction of NADH oxidation and by following the change in absorbance at 470 nm. As seen from Fig. 2, guaiacol, added after NADH oxidation, was oxidized. When the reaction mixture for NADH oxidation was preincubated in the absence of NADH for 5 min and then guaiacol was added, no oxidation of guaiacol was observed (data not shown). Thus H2O2-formation was dependent on NADH oxidation.
|
Localization of peroxidase activity
As shown above, peroxidase activity was present exclusively in the micropylar endosperm of tomato seeds during the first 2 d after imbibition (Table 1A). To confirm this further, tissue printing experiments were done. In tissue prints of seeds imbibed for 3 h, no signal of the enzyme activity appeared (data not shown). On the other hand, in 1-d-imbibed seeds or seeds of 2-d-old seedlings, the signals of the activity were clearly detected specifically in the micropylar endosperm (Fig. 3A, 1, 2). These histochemical observations are in accord with the results obtained by assaying the activities spectrophotometrically (Table 1A).
|
In the wounded lateral endosperm, the activity was detected exclusively at the wounded (cut) region (Fig. 3A, 3, 4). This is in good agreement with the observation that peroxidase activity is induced by wounding and that the wound-effect is manifested only at the wounded part (Table 1B).
Presence of H2O2 in the micropylar endosperm
Seeds of 2-d-old seedlings were laterally halved and embryo parts were removed from the halved seeds. When the de-embryonated half-seed was incubated in the presence of TMB, a blue staining appeared in the micropylar endosperm (Fig. 4A). The staining was completely precluded in the presence of an H2O2 scavenger (KI, 10 mM) or an antioxidant (ascorbic acid,1 mM) (Fig. 4B). Thus, it is clear that H2O2 accumulated in the micropylar endosperm. Since this method for H2O2 detection is based on the H2O2-dependent oxidation of TMB by peroxidases, the results do not necessarily exclude the possibility that H2O2 is also present in the regions other than the micropylar endosperm.
|
Patterns of peroxidase isozymes
Many peroxidase isozymes of tomato endosperm were visualized on an IEF gel using 4-chloro-1-naphthol as the substrate, as shown in Fig. 5. The pattern of the isozymes in the micropylar endosperm changed with time after imbibition (Fig. 5A). A band at the origin could be due to an artefact as a result of the application of tissue extracts at this area of the gel, since such artefact bands appeared wherever the samples were applied (data not shown). A few minor isozymes with pI values between 5.0 (origin) and 3.5 were observed to be present, but the reproducibility of their appearance and of their separation on IEF gels was poor. These minor isozymes were not analysed in the present study. In the micropylar endosperm of imbibed seeds for 1 d, two isozymes with pIs lower than 3.5 were the most active (dots in Fig. 5A, lane 1). In addition to these, three isozymes (pI 7.1, 7.5 and 8.0) were noticeable. In the micropylar endosperm of seeds of 2-d-old seedlings, the activities of pI 7.1 and pI 8.0 isozymes were remarkably enhanced (Fig. 5A, lane 2). The isozyme with a pI of 7.5 that was detected in 1-d-imbibed seeds disappeared in the micropylar endosperm of seeds of 2-d-imbibed seedlings, and pI 9.4 isozyme newly appeared. In the micropylar endosperm of seeds of 3-d-old seedlings, basic isozymes (pI 9.1, 9.2 and 9.3) were markedly expressed (Fig. 5A, lane 3).
|
The isozyme pattern in the lateral endosperm was studied with seeds of 3-d-old seedlings. Before this stage, peroxidase isozymes in the lateral endosperm were difficult to analyse because of their low activities. The major isozymes detectable in the lateral endosperm of seeds of 3-d-old seedlings were those with pI <3.5, 4.7 and 7.1 (Fig. 5B, lane 1). This isozyme profile of the lateral endosperm was different from that of the micropylar endosperm of 1-d-imbibed seeds (Fig. 5A, lane 1); the notable differences are that the pI 7.5 isozyme present in the micropylar endosperm was not detectable in the lateral endosperm and that the pI 4.7 isozyme detected in the lateral endosperm was not present in the micropylar endosperm. Thus, the isozyme profiles observed at the time when peroxidases began to be synthesized in the endosperm were different between the micropylar portion (1 d after imbibition) and the lateral one (3 d after imbibition). In a previous paper (Nonogaki et al., 1998), it was shown that the processes of the biochemical activation of the endosperm after imbibition are qualitatively different between the micropylar region and the lateral one. The difference in peroxidase isozyme profiles between the two endosperm regions may reflect this situation.
Effect of wounding on the expression of peroxidase isozymes
In the wounded lateral endosperm, five peroxidase isozymes represented the majority of the peroxidase activity (Fig. 5B, lane 2); two isozymes with pIs lower than 3.5 and isozymes with pIs of 3.5, 7.1 and 7.9. Among them, the pI 3.5 and 7.9 isozymes were not detectable in intact lateral endosperm. These isozymes seemed to be newly induced in response to wounding in the lateral endosperm. Anionic peroxidases have been shown to be induced in response to wounding in several plant species (Espelie et al., 1986; Kolattukudy et al., 1992; Christensen et al., 1998; Hiraga et al., 2000). On the other hand, it has also been reported that cationic peroxidases are induced by wounding (Desbiez and Boyer, 1981; Lagrimini and Rothstein, 1987; Svelheim and Robertsen, 1990; Kawaoka et al., 1994; Smith et al., 1994; Hiraga et al., 2000; Quiroga et al., 2000).
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
In tomato seeds, the embryo is surrounded by the endosperm and, when the radicle of germinating seeds penetrates the micropylar portion of the endosperm, that endosperm part is ruptured. Radicle penetration through the endosperm, therefore, results in the exposure of the inner tissues of the seed to environmental factors such as pathogenic micro-organisms; pathogens in the surrounding soil can easily enter into the inner tissues through the ruptured micropylar endosperm. There is the possibility that tomato seeds are endowed with some mechanism(s) for defence against such an invasion of pathogens. In this connection, the fact is suggestive that ß-1,3-glucanase and chitinase are expressed specifically in the micropylar endosperm in germinating and germinated tomato seeds, in spite of the absence of their substrates in the endosperm tissue (Morohashi and Matsushima, 2000; Wu et al., 2001). Since these enzymes are well known to help plants defend against fungal infection (Simmons, 1994), the induction of these enzymes in the micropylar endosperm might be a defensive mechanism of tomato seeds during and following germination.
In the present study it was shown that peroxidase activity markedly developed in the micropylar endosperm, but not in the lateral endosperm, of germinating and germinated tomato seeds (Fig. 1; Table 1). This developmental pattern is very similar to that observed with ß-1,3-glucanase and chitinase (Morohashi and Matsushima, 2000; Wu et al., 2001). There are quite a few reports showing that peroxidase is induced in response to wounding in plants (Espelie et al., 1986; Lagrimini and Rothstein, 1987; Kolattukudy et al., 1992; Siegel, 1993; Quiroga et al., 2000). The present study showed that peroxidase activity in tomato endosperm was greatly enhanced when the tissue was wounded (Table 1B; Fig. 3). Radicle penetration through the micropylar endosperm is accompanied by the rupture, or wounding, of the micropylar endopserm. It is possible that the induction of peroxidase activity in the micropylar endosperm is brought about by the radicle penetration of the tissue and is associated with defensive reactions. However, the endosperm rupturing (wounding) does not seem to be the direct cause of the induction of peroxidase activities in the micropylar endosperm, because peroxidase activity had already developed prior to radicle protrusion (Fig. 1). It has been shown that cell wall hydrolysis by endo-ß-mannanase occurs in the micropylar endosperm prior to radicle protrusion, resulting in the endosperm weakening at the site of radicle penetration (Bewley, 1997). The degradation of the cell wall may have an effect similar to wounding. Therefore, the micropylar endosperm may be wounded in a sense during germination (prior to radicle emergence). This may be the reason why the peroxidase activity develops in the micropylar endosperm prior to radicle protrusion. On the other hand, the cell wall of the lateral endosperm cells is degraded after the completion of germination (Bewley, 1997). This may result in the enhancement of peroxidase activity in the lateral endosperm at rather later stages (Fig. 1). However, this view does not explain why peroxidase activity in the micropylar endosperm is much more strongly enhanced than that in the lateral endosperm.
It is noteworthy that the isozyme pattern was different between the intact and the wounded (cut) endosperm; pI 3.5 and 7.9 isozymes that were not detected in the intact endosperm were highly expressed in the response to wounding (Fig. 5B). This may indicate that the signal which triggers the induction of peroxidase in the intact endosperm is different from that resulting from wounding (cutting).
What is the function of peroxidases expressed in the micropylar endosperm of tomato seeds? It has been reported that there is an association between peroxidase activity in plant tissues and their resistance to pathogen infection (Hammerschmidt et al., 1982; Joseph et al., 1998). There is increasing evidence suggesting that the joint exudation of H2O2 and peroxidase plays an important role in the defence system of plants against pathogens (Scott-Craig et al., 1995; Bestwick et al., 1998; Schopfer et al., 2001). In plants, the increased production of the superoxide radicals and H2O2 is a common feature of defence responses to pathogen attack (Lamb and Dixon, 1997). H2O2 released into the apoplast is directly toxic to pathogens (Wu et al., 1997; Joseph et al., 1998; De Rafael et al., 2001). It is also suggested that, in addition to direct action on the growth of pathogenic microorganisms, H2O2 (and other active oxygen species) may induce complex programmed cell death at sites attacked by pathogens, the hypersensitive reaction (Grant and Loake, 2000; Fath et al., 2001). The presence of H2O2 in the micropylar endosperm was shown by the histochemical observation (Fig. 4). Taking this into consideration, the possibility can be suggested that peroxidases induced specifically in the micropylar endosperm play a critical role, along with H2O2, in protecting the exposed inner part of the endosperm against the invasion of pathogenic organisms.
As shown above (Fig. 2), tomato peroxidase catalyses the synthesis of H2O2 at the expense of NADH. However, there is no evidence that this oxidase activity is physiologically significant in the tissues. Recently, a model of pH-dependent generation of H2O2 by a cell-wall peroxidase has been proposed; although the reductant of the peroxidase in this model has not been identified, NADH is not likely to be the reductant (Bolwell et al., 1995; Wojtaszek, 1997). There are also reports indicating that H2O2 generation can be mediated by enzymes other than peroxidase (Bolwell and Wojtaszek, 1997; Frahry and Schopfer, 1998).
It is noteworthy that peroxidase activity develops in the micropylar endosperm of tomato seeds prior to radicle protrusion, i.e. prior to the exposure of the inner tissues to a hazardous environment. It has recently been shown that active oxygen species are released to the apoplast from embryos and aleurone layers of radish seeds prior to radicle emergence (Schopfer et al., 2001). It is, therefore, conceivable that peroxidase expression accompanied by H2O2 generation is carried out in anticipation of radicle penetration through the endosperm and gives the seed a pre-emptive protective mechanism for survival in hazardous environments. The fact that peroxidase activity begins to develop in germinating seeds without any environmental cues such as pathogen attack or wounding, may indicate that peroxidase expression may be a developmentally regulated phenomenon and a constitutive biochemical process in germinating seeds. It has recently been shown that roots of soybean, sunflower and maize seedlings are able to produce H2O2 in the absence of pathogen attack (Frahry and Schopfer, 1998).
The developmental stage (days 1 and 2) when germination completes and the radicle ruptures the endosperm, is thought to represent the time most sensitive for pathogen infection, as pointed out by Schopfer et al. (2001). At later stages, the upper part of the seedling (hypocotyl and cotyledon) appears above the soil and the seed part (endosperm remnants and testa) is shed from the seedling. However, peroxidase activity in the micropylar endosperm of tomato seeds continued to increase markedly until day 3 and reached a level more than 200-fold higher than that at day 1. Is it necessary for peroxidase activity to continue to increase in the micropylar endosperm until rather later stages? Furthermore, pI 9.1, 9.2 and 9.3 isozymes were newly expressed in the micropylar endosperm between days 2 and 3 (Fig. 5A). If they play a defensive role, why are they not expressed at earlier stages? It is difficult to explain the reason from the viewpoint of defensive responses why the pattern of peroxidase isozymes in the micropylar endosperm changes so markedly with time (Fig. 5A). These problems remain to be solved in future. It is necessary to isolate each isozyme, characterize it and identify its function.
Lignification or suberization is thought to be an important wound-healing mechanism (Espelie et al., 1986; Angelini et al., 1993). Peroxidase has been proposed to play a role in the oxidation of monoglinols prior to their polymerization during lignin formation, although other oxidases may also be involved (Whetten et al., 1998). Peroxidases are also suggested to be critical in the polymerization of aromatic monomers to suberin (Kolattukudy, 1981). Lignin formation in the endosperm was examined histochemically by the phloroglucinol–HCl method (Barcelo, 1998). No signal for the presence of lignin in the micropylar endosperm was obtained.
Further investigations are needed in order to elucidate the physiological roles of peroxidases that develop in the endosperm of germinating and germinated tomato seeds.
|
Acknowledgements |
|---|
The author thanks Dr Yasuko Kaneko for help in microscopy observations and Ms Sachiko Morohashi for practical assistance.
|
References |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Angelini R, Bragaloni M, Federico R, Infantino A, Porta-Puglia A. 1993. Involvement of polyamines, diamine oxidase and peroxidase in resistance of chickpea to Ascochyta rabiei. Journal of Plant Physiology 142, 704–709.[Web of Science]
Barcelo AR. 1998. Hydrogen peroxide production is a general property of the lignifying xylem from vascular plants. Annals of Botany 82, 97–103.
Bestwick CS, Brown IR, Manfield JW. 1998. Localized changes in peroxidase activity accompany hydrogen peroxide generation during the development of a non-host hypersensitive reaction in lettuce. Plant Physiology 118, 1067–1078.
Bewley JD. 1997. Breaking down the walls—a role for endo-ß-mannanase in release from seed dormancy? Trends in Plant Science 2, 464–469.[Web of Science]
Bolwell GP, Butt VS, Davies DR, Zimmerlin A. 1995. The origin of the oxidative burst in plants. Free Radical Research 23, 517–532.[Web of Science][Medline]
Bolwell GP, Wojtaszek P. 1997. Mechanisms for the generation of reactive oxygen species in plant defense – a broad perspective. Physiological and Molecular Plant Pathology 51, 347–366.
Christensen JH, Bauw G, Welinder KG, van Montagu M, Boerjan W. 1998. Purification characterization of peroxidases correlated with lignification in poplar xylem. Plant Physiology 118, 125–135.
De Rafael MA, Valle T, Babiano MJ, Corchete P. 2001. Correlation of resistance and H2O2 production in Ulmus pumila and Ulmus campestris cell suspension cultures inoculated with Ophiostoma novo-ulmi. Physiologia Plantarum 111, 512–518.[Medline]
Derckel JP, Audran JC, Haye B, Lambert B, Legendre L. 1998. Characterization, induction by wounding and salicylic acid, and activity against Botrytis cinerea of chitinases and ß-1,3-glucanases of ripening grape berries. Physiologia Plantarum 104, 56–64.
Desbiez MO, Boyer N. 1981. Hypocotyl growth and peroxidases of Bidens pilosus. Effect of cotyledonary prickings and lithium pretreatment. Plant Physiology 68, 41–43.
Espelie KE, Franceschi VR, Kolattukudy PE. 1986. Immunocytochemical localization and time-course appearance of an anionic peroxidase associated with suberization in wound-healing potato tuber tissue. Plant Physiology 81, 487–492.
Fath A, Bethke PC, Jones RL. 2001. Enzymes that scavenge reactive oxygen species are down-regulated prior to gibberellic acid-induced programmed cell death in barley aleurone. Plant Physiology 126, 156–166.
Frahry G, Schopfer P. 1998. Hydrogen peroxide production by roots and its stimulation by exogenous NADH. Physiologia Plantarum 103, 395–404.
Grant JJ, Loake GL. 2000. Role of reactive oxygen intermediates and cognate redox signaling in disease resistance. Plant Physiology 124, 21–29.
Halliwell B. 1978. Lignin synthesis: the generation of hydrogen peroxide and superoxide by horseradish peroxidase and its stimulation by manganese (II) and phenols. Planta 140, 81–88.[Web of Science]
Hammerschmidt R, Nuckles EM, Kuc J. 1982. Association of enhanced peroxidase activity with induced systemic resistance of cucumber to Colletotrichum lagenarium. Physiogical Plant Pathology 20, 73–82.
Hiraga S, Ito H, Sasaki K, Yamakawa H, Mitsuhara I, Toshima H, Matsui H, Homma M, Ohashi Y. 2000. Wound-induced expression of a tobacco peroxidase is not enhanced by ethephon and suppressed by methyl jasmonate and coronatine. Plant and Cell Physiology 41, 165–170.
Joseph LM, Koon TT, Man WS. 1998. Antifungal effects of hydrogen peroxide and peroxidase on spore germination and mycelial growth of Pseudocercospora species. Canadian Journal of Botany 76, 2119–2124.
Kato M, Hayakawa Y, Hyodo H, Ikoma Y, Yano M. 2000. Wound-induced ethylene synthesis and expression and formation of 1-aminocyclopropane-1-carboxylate (ACC) synthase, ACC oxidase, phenylalanine ammonia-lyase, and peroxidase in wounded mesocarp tissue of Cucurbita maxima. Plant and Cell Physiology 41, 440–447.
Kawaoka A, Kawamoto T, Ohta H, Sekine M, Takano M, Shinmyo A. 1994. Wound-induced expression of horseradish peroxidase. Plant Cell Reports 13, 149–154.
Kolattukudy PE. 1981. Structure, biosynthesis and biodegradation of cutin and suberin. Annual Review of Plant Physiology 32, 539–567.[Web of Science]
Kolattukudy PE, Mohan R, Bajar MA, Serf BA. 1992. Plant peroxidase gene expression and function. Biochemical Society Transactions 20, 333–337.
Lagrimini LM, Rothstein S. 1987. Tissue specificity of tobacco peroxidase isozymes and their induction by wounding and tobacco mosaic virus infection. Plant Physiology 84, 438–442.
Lamb C, Dixon R. 1997. The oxidative burst in plant disease resistance. Annual Review of Plant Physiology and Plant Molecular Biology 48, 251–275.[Web of Science]
Leubner-Metzger G, Frundt C, Vogeli-Lange R, Meins Jr F. 1995. Class I ß-1,3-glucanase in the endosperm of tobacco during germination. Plant Physiology 109, 751–759.[Abstract]
Maeder M, Amberg-Fisher V. 1982. Role of peroxidase in lignification of tobacco cells. I. Oxidation of nicotinamide adenine dinucleotide and formation of hydrogen peroxide by cell wall peroxidases. Plant Physiology 70, 1128–1131.
Maeder M, Ungemach J, Schloss P. 1980. The role of peroxidase isozyme groups of Nicotiana tabacum in hydrogen peroxide formation. Planta 147, 467–470.
Mauch F, Hadwiger LA, Boller T. 1988. Antifungal hydrolases in pea tissue. I. Purification and characterization of two chitinases and two ß-1,3-glucanases differentially regulated during development and in response to fungal infection. Plant Physiology 87, 325–333.
Morohashi Y, Matsushima H. 2000. Development of ß-1,3-glucanase activity in germinated tomato seeds. Journal of Experimental Botany 51, 1381–1387.
Nonogaki H, Matsushima H, Morohashi Y. 1992. Galactomannan hydrolysing activity develops during priming in the micropylar endosperm tip of tomato seeds. Physiologia Plantarum 85, 167–172.
Nonogaki H, Nomaguchi M, Okumoto N, Kaneko Y, Matsushima H, Morohashi Y. 1998. Temporal and spatial pattern of the biochemical activation of the endosperm during and following imbibition of tomato seeds. Physiologia Plantarum 102, 236–242.
Petruzzelli L, Kunz C, Waldvogel R, Meins Jr F, Leubner-Metzger G. 1999. Distinct ethylene- and tissue-specific regulation of ß-1,3-glucanases and chitinase during pea seed germination. Planta 209, 195–201.[Web of Science][Medline]
Quiroga M, Guerrero G, Botella MA, Barcelo A, Amaya I, Medina MI, Alonso FJ, de Forchetti SM, Tigier H, Valpuesta V. 2000. A tomato peroxidase involved in the synthesis of lignin and suberin. Plant Physiology 122, 1119–1127.
Schopfer P, Plachy C, Frahry G. 2001. Release of reactive oxygen intermediates (superoxide radicals, hydrogen peroxide, and hydroxyl radicals) and peroxidase in germinating radish seeds controlled by light, gibberellin, and abscisic acid. Plant Physiology 125, 1591–1602.
Scott-Craig JS, Kerby KB, Stein BD, Sommerville CS. 1995. Expression of an extracellular peroxidase that is induced in barley by the powdery mildew pathogen (Erysiphe graminis f.sp. hrodei). Physiological and Molecular Plant Pathology 47, 407–418.
Siegel BZ. 1993. Plant peroxidases—an organismic perspective. Plant Growth Regulation 12, 303–312.
Simmons CR. 1994. The physiology and molecular biology of plant 1,3-ß-glucanases and 1,3-;1,4-ß-glucanases. Critical Reviews in Plant Sciences13, 325–387.
Smith CG, Rodgers M, Zimmerlin A, Ferdinando D, Bolwell GP. 1994. Tissue and subcellular immunolocalization of enzymes of lignin synthesis in differentiating and wounded hypocotyl tissue of French bean (Phaseolus vulgaris L.). Planta 192, 155–164.
Svelheim F, Robertsen B. 1990. Induction of peroxidases in cucumber hypocotyls by wounding and fungal infection. Physiologia Plantarum 78, 261–267.
Vogeli-Lange R, Frundt C, Hart CM, Beffa R, Nage F, Meins Jr F. 1994. Evidence for a role of ß-1,3-glucanase in dicot seed germination. The Plant Journal 5, 273–278.
Whetten RW, MacKay JJ, Sederoff RR. 1998. Recent advances in understanding lignin biosynthesis. Annual Review of Plant Physiology and Plant Molecular Biology 49, 585–609.[Web of Science]
Wojtaszek P. 1997. Oxidative burst: an early plant response to pathogen infection. Biochemical Journal 322, 681–692.
Wu CT, Leubner-Metzger G, Meins Jr F, Bradford KJ. 2001. Class I ß-1,3-glucanase and chitinase are expressed in the micropylar endosperm of tomato seeds prior to radicle emergence. Plant Physiology 126, 1299–1313.
Wu G, Shortt BJ, Lawrence EB, Leon J, Fitzsimmons KC, Levine EB, Raskin I, Shah DM. 1997. Activation of host defense mechanisms by elvated production of H2O2 in transgenic plants. Plant Physiology 115, 427–435.[Abstract]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  A. Linkies, U. Schuster-Sherpa, S. Tintelnot, G. Leubner-Metzger, and K. Muller Peroxidases identified in a subtractive cDNA library approach show tissue-specific transcript abundance and enzyme activity during seed germination of Lepidium sativum J. Exp. Bot., October 30, 2009; (2009) erp318v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Fan, H. Wang, D. Feng, B. Liu, H. Liu, and J. Wang Molecular Characterization of Plantain Class I Chitinase Gene and its Expression in Response to Infection by Gloeosporium musarum Cke and Massee and other Abiotic Stimuli J. Biochem., November 1, 2007; 142(5): 561 - 570. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C.-T. Wu and K. J. Bradford Class I Chitinase and {beta}-1,3-Glucanase Are Differentially Regulated by Wounding, Methyl Jasmonate, Ethylene, and Gibberellin in Tomato Seeds and Leaves Plant Physiology, September 1, 2003; 133(1): 263 - 273. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||