Possible role of catalase in post-dormancy bud break in grapevines

Changes in the activity of catalase (Cat) and in the levels of H2O2 were followed throughout dormancy in buds of grapevines (Vitis vinifera L.). In grapevines grown in the Elqui valley in Chile, a region with warm-winters, the activity of Cat increased during the recess period of buds, reaching a maximum and thereafter decreased to less than one third of its maximal activity. Three isoforms of Cat were detected in extracts of buds by native PAGE analysis, and the extracted activity was inhibited competitively by hydrogen cyanamide (HC), a potent bud-break agent. Furthermore, HC applications to field-grown grapevines in addition to the expected effect on advancing bud break, reduced the Cat activity during bud dormancy. Similar reductions were observed during dormancy in buds of grapevines grown in the Central valley in Chile, a region with temperate winters, suggesting that HC and winter chilling inhibits the activity of the main H2O2 degrading enzyme in grape buds. A transient rise in H2O2 levels preceded the release of buds from endodormancy, moreover, the peak of H2O2 and the onset of bud break occurred earlier in HC treated than in control grapevines, suggesting the participation of H2O2 as a signal molecule in the release of endodormancy in grape buds. The relationship between Cat inhibition, rise in H2O2 levels and initiation of bud break are discussed.     

Involvement of calcium signalling in dormancy release of grape buds

Artificial induction of grape bud dormancy release by hydrogen cyanamide (HC) serves as a reliable model system to explore the events occurring shortly after the induction of dormancy release. Recently, a group of genes with remarkable differences in expression level between HC-treated and control buds was identified. The identification of several calcium signalling-related genes within that group raised the hypothesis of the involvement of Ca(2+) signalling in grape bud dormancy release. Therefore, the effects of HC treatment on the expression profiles of several calcium sensors, the effect of the plasma membrane calcium channel blocker LaCl(3) and the calcium chelator EGTA on HC-induced and chilling-induced bud-break, and the effect of HC application on calcium-dependent protein phosphorylation activities in the bud tissue were studied. Here the HC-induced expression of Ca(2+)-ATPase is described, indicating that this treatment might evoke an increase in [Ca(2+)]cyt. Similar induction was confirmed for calmodulin, calmodulin-binding protein, and calcium-dependent protein kinase (CDPK). Both LaCl(3) and EGTA blocked the inducing effect of HC on bud-break, and their inhibitory effects were removed by supplying exogenous Ca(2+). Calcium-dependent histone phosphorylation was up to 70% higher in HC-treated buds. Endogenous protein phosphorylation assays detected four proteins exhibiting increased phosphorylation following HC treatment, of which two were phosphorylated in a calcium-dependent manner. One of these, a 47 kDa protein, presented strong and Ca(2+)-dependent phosphorylation only in HC-treated buds. The potential role of CDPK in the phosphorylation of this protein was supported by an immunoprecipitation assay. The data suggest, for the first time, that calcium signalling is involved in the mechanism of bud dormancy release.     

高温和单氰胺对油桃休眠花芽呼吸代谢的影响

以3年生盆栽‘曙光’油桃为材料,研究油桃自然休眠过程中50℃高温和单氰胺对花芽呼吸代谢的影响.结果表明:高温和单氰胺均可以打破油桃的自然休眠,导致休眠花芽呼吸代谢显著下降,其呼吸代谢的衰减可持续数小时.主要呼吸途径三羧酸循环(TCA)和磷酸戊糖途径(PPP)的运行均受到影响.未经破眠处理的花芽TCA和PPP均呈衰减趋势,而高温和单氰胺诱导了早期呼吸衰减结束后PPP的迅速活化.高温还表现出对TCA恢复的诱导,而单氰胺在96 h内未表现出这种作用.在高温和单氰胺打破自然休眠的机制中,呼吸衰减和随后出现的PPP活化可能是重要的组成部分.     

Abscisic acid catabolism enhances dormancy release of grapevine buds

The molecular mechanism regulating dormancy release in grapevine buds is as yet unclear. It was formerly proposed that dormancy is maintained by abscisic acid (ABA)‐mediated repression of bud–meristem activity and that removal of this repression triggers dormancy release. It was also proposed that such removal of repression may be achieved via natural or artificial up‐regulation of VvA8H‐CYP707A4, which encodes ABA 8′‐hydroxylase, and is the most highly expressed paralog in grapevine buds. The current study further examines these assumptions, and its experiments reveal that (a) hypoxia and ethylene, stimuli of bud dormancy release, enhance expression of VvA8H‐CYP707A4 within grape buds, (b) the VvA8H‐CYP707A4 protein accumulates during the natural transition to the dormancy release stage, and (c) transgenic vines overexpressing VvA8H‐CYP707A4 exhibit increased ABA catabolism and significant enhancement of bud break in controlled and natural environments and longer basal summer laterals. The results suggest that VvA8H‐CYP707A4 functions as an ABA degrading enzyme, and are consistent with a model in which the VvA8H‐CYP707A4 level in the bud is up‐regulated by natural and artificial bud break stimuli, which leads to increased ABA degradation capacity, removal of endogenous ABA‐mediated repression, and enhanced regrowth. Interestingly, it also hints at sharing of regulatory steps between latent and lateral bud outgrowth.     

Influence of different temperatures on bud break and 1-aminocyclopropane-1-carboxylic acid content of young peach trees

Effects of different temperatures on bud break and 1-aminocyclopropane-1-carboxylic acid (ACC) content were determined by using potted two-year-old ‘Akatsuki’ peach trees. One group of trees were subjected to 1°C for four weeks and then transferred to a growth chamber at 24°C, while the other was kept at 24 °C throughout the experiment. After four-week temperature treatments floral and vegetative bud break were evaluated weekly and bud break percentage was calculated. Bud break was greater under 1 °C than 24 °C in both November and December. The time required to release buds from dormancy was shorter in December than November. In November ACC content in peach buds increased after one and two weeks, then decreased in the forth week under both treatments. However, in December ACC content after two and four weeks showed a similar trend under 1 °C and a reverse trend under 24 °C. It was higher under low temperature treatment. These data indicate that chilling requirements for bud break of peach seems to be associated with the promotion of ethylene biosynthesis caused by low temperature stress.     

Subtle proteome differences identified between post-dormant vegetative and floral peach buds

Proper development of deciduous tree species, including peach, is accomplished through an annual growth cycle. Freezing avoidance during winter is necessary for tree survival and is achieved by the enclosure of meristems in floral and vegetative buds. To elucidate the role of developmentally regulated protein networks in bud break, proteins of the two bud-types were extracted and analyzed by two-dimensional gel electrophoresis (2-DE). Of the 1107 protein spots that were picked, 475 were identified and annotated assembling the peach bud proteome reference map. The majority of these proteins are involved in stress-response, detoxification, defense, carbohydrate metabolism and energy production. The protein profiles of both bud-types bear high similarity, whereas only 11 proteins were differentially expressed. These proteins were mainly involved in carbon-nitrogen homeostasis/metabolism and certain developmental processes to sustain rapid growth of the newly emerging organs. Among these are enzymes that differentially regulate the levels of H(2)O(2) between floral and vegetative buds, potentially promoting sequential bud-break. Distinct Nucleoside Diphosphate Kinase (NDPK) variants in floral and vegetative buds were detected suggesting the potential role of NDPKs in H(2)O(2)-mediated signaling for post-dormant bud break. This study provides data towards a better understanding of dormancy release and bud break.     

Co-ordinated gene expression during phases of dormancy release in raspberry (Rubus idaeus L.) buds

Bud break in raspberry (Rubus idaeus L.) is often poor and uneven, with many of the subapical buds remaining in a dormant state. In order to determine the dormancy status of raspberry buds, an empirical measure of bud burst in a growth-permissive environment following exposure to chilling (4 degrees C cold storage) was developed. For cv. Glen Ample, percentage bud burst in intact canes and isolated nodes was recorded after 14 d. Isolated nodes (a measure of endodormancy) achieved 100% bud burst after approximately 1500 h chilling whereas buds on intact plants (combined endo- and paradormancy) required an additional 1000 h chilling. A microarray approach was used to follow changes in gene expression that occurred during dormancy transition. The probes for the microarrays were obtained from endodormant and paradormant raspberry bud cDNA libraries. The expression profiles of 5300 clones from these libraries were subjected to principal component analysis to determine the most significant expression patterns. Sequence analysis of these clones, in many cases, enabled their functional categorization and the development of hypotheses concerning the mechanisms of bud dormancy release. Thus a set of novel candidates for key dormancy-related genes from raspberry buds have been identified. Bud dormancy is fundamental to the study of plant developmental processes and, in addition, its regulation is of significant economic importance to fruit and horticultural industries.     

Interaction of endodormancy and paradormancy in raspberry (Rubus idaeus L.)

Bud break in protected Northern European raspberry crops is often poor and uneven with many of the sub-apical buds remaining in a dormant state. In order to improve bud break and therefore yields, the mechanism controlling bud dormancy must be determined. Canes of the biennial cultivar ‘Glen Moy’ were forced as isolated single nodes, trisections, or as intact canes after different lengths of cold storage chill unit (CU) accumulation in order to determine whether the buds were in an endodormant or paradormant state. The results showed that buds on the lower parts of the intact canes remained in a dormant state long after buds from higher up the intact cane and also the single nodes from all parts of the cane had emerged from the deepest phase of endodormancy. This would imply that these buds were being held in a paradormant state until large amounts of chilling units (>1000 CU) had been accumulated. The trisected cane portions revealed almost no significant differences in bud break levels throughout the experiment when compared with the single nodes. This suggests that removal of the apical part of the cane would be effective in improving bud break by reducing the paradormant condition. A period of secondary dormancy was also observed in the intact canes which may also exacerbate the poor bud break observed in protected crops. This was not seen in the single nodes or the trisected canes which indicates that treatments which reduce paradormancy may also minimise the risk of secondary dormancy. By identifying the phase of bud dormancy which causes poor bud break, attention can now be focused on methods which overcome paradormancy in protected crops. Such methods might include tipping (removal of the cane apex), horizontal training methods, more efficient chilling methods, and chemical treatments.     

Exogenous dormancy-breaking substances positively change endogenous phytohormones and amino acids during dormancy release in ‘Anna’ apple trees

A 2-season trial was conducted to verify the effects of foliar applications of some dormancy-breaking substances (DBS) on dormancy release in buds of ‘Anna’ apple (Malus sylvestris, Mill) trees, as well as on metabolic changes in the contents of phytohormones, proline and arginine in buds during their release from dormancy. The efficiency of early bud break induced by Dormex?, potassium nitrate, mineral oil, calcium nitrate and thiourea was noticed in varying degrees. Although Dormex? was distinguished, all DBS hastened bud break, shortened flowering duration, improved bud break% and fruit-set%, increased the contents of gibberellic acid, indole-3-acetic acid, proline and arginine, but reduced abscisic acid content in buds as compared to the control. These results were positively reflected in the final tree yield. Accordingly, it is concluded that the use of Dormex?, at a rate of 4 %, could be recommended for reaching bud break as early as possible and improving ‘Anna’ apple tree yield under the short winters in Egypt and similar regions by regulating the contents of proline, arginine and phytohormones in buds.     

Anatomical and Histochemical Changes of Norway Spruce Buds Induced by Simulated Acid Rain

The study was focused on changes of anatomical and histochemical parameters of buds of 4-year-old Norway spruce (Picea abies L. Karst) trees subjected to simulated acid rain (SAR). Solutions of pH 2.9 and 3.9 were applied by spraying on shoot and/or by watering for two years. No macroscopic changes of buds or needles were observed in connection with SAR application and the only induced change was 2-week earlier onset of bud break in all treated variants compared to the control. Two-year treatment caused decrease in number of leaf primordia and increase in number of living bud scales in treated dormant buds while these parameters remained unchanged in the control buds. Treatments with solution of pH 2.9 caused decrease of flatness of bud apical meristem during the vegetative season. Increased activity of non-specific esterase in treated buds occurred during dormancy and bud break and the enhanced accumulation of phenolic compounds was detected at the beginning of shoot growth. Changes in histochemical parameters of bud tissues were induced mainly by spraying of shoots and can thus be qualified as primary damage.     

Is GABA-shunt functional in endodormant grapevine buds under respiratory stress?

It has been suggested that a respiratory stress is part of the mechanism through which the dormancy-breaking compounds, hydrogen cyanamide (HC) and sodium azide, induce the release of buds from the endodormancy (ED) in grapevines. The accumulation of metabolites like succinate, alanine (Ala) and γ-amino butyric acid (GABA), together with the activation of the GABA-shunt pathway, is a general feature of plants in response to oxygen deprivation and to respiratory stress. Unexpectedly, in a previous study, we found that GABA applied exogenously to grapevine buds, down-regulated the expression of most genes encoding for antioxidant enzymes, suggesting that its accumulation under respiratory stress conditions could be deleterious for the bud. In order to analyze whether GABA accumulates under respiratory stress conditions in grapevine buds, we analysed in this study, the effect of hypoxia, the respiration inhibitor KCN and the dormancy breaker compound HC, on the level of GABA, and on the expression levels of the GABA-shunt genes (VvGAD, VvGABA-T, VvSSADH). Additionally, genes from the Ala fermentative pathway (VvAlaAT, VvAspAT) were also analysed. The results revealed that although the three treatments mentioned above, up-regulated the expression of VvGAD1, the content of GABA remained constant, while Ala content increased. The lack of GABA accumulation under respiratory stress is an important physiological fact in grapevine buds, since it avoids the down-regulation of antioxidant genes, and promotes the incorporation of succinate into the TCA cycle, a fact that would be important in the release of buds from the ED.     

Spatial and temporal changes in multiple hormone groups during lateral bud release shortly following apex decapitation of chickpea (Cicer arietinum) seedlings

Although the co-ordination of promotive root-sourced cytokinin (CK) and inhibitory shoot apex-sourced auxin (IAA) is central to all current models on lateral bud dormancy release, control by those hormones alone has appeared inadequate in many studies. Thus it was hypothesized that the IAA : CK model is the central control but that it must be considered within the relevant timeframe leading to lateral bud release and against a backdrop of interactions with other hormone groups. Therefore, IAA and a wide survey of cytokinins (CKs), were examined along with abscisic acid (ABA) and polyamines (PAs) in released buds, tissue surrounding buds and xylem sap at 1 and 4 h after apex removal, when lateral buds of chickpea are known to break dormancy. Three potential lateral bud growth inhibitors, IAA, ABA and cis -zeatin 9-riboside (ZR), declined sharply in the released buds and xylem following decapitation. This is in contrast to potential dormancy breaking CKs like trans -ZR and trans -zeantin 9-riboside 5'phosphate (ZRMP), which represented the strongest correlative changes by increasing 3.5-fold in xylem sap and 22-fold in buds. PAs had not changed significantly in buds or other tissues after 4 h, so they were not directly involved in the breaking of bud dormancy. Results from the xylem and surrounding tissues indicated that bud CK increases resulted from a combination synthesis in the bud and selective loading of CK nucleotides into the xylem from the root.     

Alterations in the pattern of peroxidase isoenzymes and transient increases in its activity and in H2O2 levels take place during the dormancy cycle of grapevine buds: the effect of hydrogen cyanamide

Polymorphism of peroxidase (Px) and changes in its activity and in H₂O₂ content were studied in buds of grapevine during dormancy. Three isoforms of Px were detected in bud-extracts, two basic and one acidic, however, the pattern of Px isoenzyme changed with the progress of dormancy. Thus, basic Px isoenzymes disappeared from extracts previous to the onset of bud-break, while acidic isoenzymes remained relatively unaltered throughout the whole dormancy period. Furthermore, transient increases in the activity of Px and in the content of H₂O₂ occurred previous to endodormancy release, when buds were fully dormant. Hydrogen cyanamide (H₂CN₂), a potent bud breaking agent in grapevines advanced as expected bud-break, but also advanced the occurrence of Px and H₂O₂ peaks and the changes in Px isoenzymes pattern. The results suggests that H₂O₂ could function as a signalling molecule inducing endodormancy release, and changes in Px polymorphism could be a useful marker to study endo/ecodormancy phase transition in buds of grapevines.     

Release of apical dominance in potato tuber is accompanied by programmed cell death in the apical bud meristem

Potato (Solanum tuberosum) tuber, a swollen underground stem, is used as a model system for the study of dormancy release and sprouting. Natural dormancy release, at room temperature, is initiated by tuber apical bud meristem (TAB-meristem) sprouting characterized by apical dominance (AD). Dormancy is shortened by treatments such as bromoethane (BE), which mimics the phenotype of dormancy release in cold storage by inducing early sprouting of several buds simultaneously. We studied the mechanisms governing TAB-meristem dominance release. TAB-meristem decapitation resulted in the development of increasing numbers of axillary buds with time in storage, suggesting the need for autonomous dormancy release of each bud prior to control by the apical bud. Hallmarks of programmed cell death (PCD) were identified in the TAB-meristems during normal growth, and these were more extensive when AD was lost following either extended cold storage or BE treatment. Hallmarks included DNA fragmentation, induced gene expression of vacuolar processing enzyme1 (VPE1), and elevated VPE activity. VPE1 protein was semipurified from BE-treated apical buds, and its endogenous activity was fully inhibited by a cysteinyl aspartate-specific protease-1-specific inhibitor N-Acetyl-Tyr-Val-Ala-Asp-CHO (Ac-YVAD-CHO). Transmission electron microscopy further revealed PCD-related structural alterations in the TAB-meristem of BE-treated tubers: a knob-like body in the vacuole, development of cytoplasmic vesicles, and budding-like nuclear segmentations. Treatment of tubers with BE and then VPE inhibitor induced faster growth and recovered AD in detached and nondetached apical buds, respectively. We hypothesize that PCD occurrence is associated with the weakening of tuber AD, allowing early sprouting of mature lateral buds.