Sumiko Sugaya

Salinity induces carbohydrate accumulation and sugar-regulated starch biosynthetic genes in tomato (Solanum lycopersicum L. cv. 'Micro-Tom') fruits in an ABA- and osmotic stress-independent manner.

Salinity stress enhances sugar accumulation in tomato (Solanum lycopersicum) fruits. To elucidate the mechanisms underlying this phenomenon, the transport of carbohydrates into tomato fruits and the regulation of starch synthesis during fruit development in tomato plants cv. ‘Micro-Tom’ exposed to high levels of salinity stress were examined. Growth with 160 mM NaCl doubled starch accumulation in tomato fruits compared to control plants during the early stages of development, and soluble sugars increased as the fruit matured. Tracer analysis with 13C confirmed that elevated carbohydrate accumulation in fruits exposed to salinity stress was confined to the early development stages and did not occur after ripening. Salinity stress also up-regulated sucrose transporter expression in source leaves and increased activity of ADP-glucose pyrophosphorylase (AGPase) in fruits during the early development stages. The results indicate that salinity stress enhanced carbohydrate accumulation as starch during the early development stages and it is responsible for the increase in soluble sugars in ripe fruit. Quantitative RT-PCR analyses of salinity-stressed plants showed that the AGPase-encoding genes, AgpL1 and AgpS1 were up-regulated in developing fruits, and AgpL1 was obviously up-regulated by sugar at the transcriptional level but not by abscisic acid and osmotic stress. These results indicate AgpL1 and AgpS1 are involved in the promotion of starch biosynthesis under the salinity stress in ABA- and osmotic stress-independent manners. These two genes are differentially regulated at the transcriptional level, and AgpL1 is suggested to play a regulatory role in this event.

Differential adaptation of high- and low-chill dormant peaches in winter through aquaporin gene expression and soluble sugar content

Plants have their own mechanisms for overcoming various stresses. In cold regions, plants are subject to stress and must enter an inherent dormancy, through several complex mechanisms, if they are to continue to exist. In winter, regulation of tonoplast and plasma membrane aquaporin genes differed in the bud cushions of the high-chill peach (Prunus persica L. Batsch) cv. Kansuke Hakuto and the low-chill peach cv. Coral. In December and January, when the temperature was lowest (around 2°C), the increased expression of Pp-γTIP1 and Pp-PIP1 seen in the bud cushions of Kansuke Hakuto may have been related to the concomitant high-soluble sugar content of the cushions of this cultivar. This relationship may have made the cells highly stable and relatively unaffected by low-temperature stress owing to the presence of “glasses” that prevented ice nucleation. However, a simpler form of cold protection regulation seemed to occur in Coral, in which there was no winter increase in Pp-γTIP1 and Pp-PIP1 mRNA and a slow decline in total soluble sugar content in December and January. These results suggested that Pp-γTIP1 and Pp-PIP1, respectively, play important roles in intra- and intercellular membrane transport, enhancing cold resistance in the bud cushions of high-chill cultivars. In addition, Pp-δTIP1 and Pp-PIP2 mRNA increased at the end of endodormancy in both cultivars. This change may be induced by endodormancy-release signals and the resumption of bud activity in both cultivars.

Identification of a soluble auxin-binding protein as a glutathione-dependent formaldehyde dehydrogenase

Antibodies against the putative auxin-binding site of an ER auxin-binding protein from maize coleoptiles were raised by injecting rabbits with a synthetic oligopeptide (Ile-His-Arg-His-Ser-Cys-Glu) conjugated to hemocyanin from the Keyhole limpet. Soluble auxin-binding proteins were isolated from an extract of etiolated mung bean hypocotyls by affinity chromatography on 2,4-dichlorophenoxyacetic acid-linked (2,4-D-linked) Sepharose 4B. The antiserum against the oligopeptide recognized a specific 40 kDa polypeptide among auxin-binding proteins from mung bean hypocotyls. An auxin-binding protein of 40 kDa was then purified by several column-chromatographic steps. The apparent molecular mass of the protein was estimated to be 80 kDa by gel-filtration and 40 kDa by SDS-PAGE. We designated this protein ABP40. The dissociation constant of purified ABP40 for [14C]-2,4-D was calculated to be 1 × 10−5 M. Binding of [14C]-2,4-D was completely inhibited by p-chlorophenoxyisobutyric acid (PCIB) and weakly inhibited by indole-3-acetic acid (IAA) and naphthalene-1-acetic acid (NAA). Benzoic acid and tryptophan had no effect on the binding. The partial amino acid sequences of ABP40, obtained after chemical cleavage by CNBr, revealed high homology to glutathione-dependent formaldehyde dehydrogenase (FDH; EC 1. 2. 1. 1). Moreover, the purified ABP40 had FDH activity.

Effect of oscillating temperature on the expression of two aquaporin genes (Pp-×TIP1,Pp-PIP2) involved in regulating intercellular water status in flower buds of peach

Summary We studied the effects of temperature changes on the water status of floral buds in peach during ecodormancy by an analysis of aquaporin (AQP) gene expression and magnetic resonance imaging of the upper part of the bud, the bud base, the bud trace, and the bud cushion. Expression levels of mRNAs of the water channel genes, Pp- TIP1 and Pp-PIP2, in the tonoplast and plasma membrane reflected the temperature oscillations: high temperatures increased mRNA levels and low temperatures decreased them, irrespective of the duration of either treatment.The T2 relaxation time of the buds, especially in the floral primordia, was significantly longer under oscillating temperature conditions than under a consistently high temperature. The period of high-temperature during the oscillating temperature regime accelerated water flow in the bud, but delayed bud growth. Disruption of the water balance by excessive water in the primordia under oscillating temperatures may be one reason for the delay in bud growth.

Carbohydrate metabolism in ‘Housui’ Japanese pear floral buds exposed to different temperatures during endodormancy

To elucidate the effects of temperature on carbohydrate metabolism under mild winter conditions, Japanese pear ( Pyrus pyrifolia Nakai) shoots were exposed to temperatures of 0, 6 and 12 °C for 600 hours during endodormancy, and subsequent bud dormancy release and carbohydrate metabolism were examined. Lateral floral buds were collected after 0, 200, 400, and 600 hours under the treatments, and after the accumulation of 2000, 4000, 6000, and 8000 growing degree hours (GDH) under heat accumulation, for carbohydrate and enzyme activities analysis. The bud burst was earliest for the 12 °C treatment, followed by the 6 and 0 °C treatments. Sucrose concentration under heat accumulation decreased early in buds from the 12 °C treatment. The sorbitol and sucrose concentration in the buds increased under 0 °C during the endodormancy. In contrast, glucose and fructose concentrations tended to increase under heat accumulation. Sucrose synthase (SS) activity increased early under heat accumulation in buds from the 6 and 12 °C treatments, but in 0 °C buds it increased more slowly, from 2000 GDH onwards. Similarly, soluble acid invertase (AI) activity increased markedly in all treatments from the middle of the heat accumulation period. We therefore suggest that a temperature of 12 °C during endodormancy may advance bud burst as the result of earlier conversion of sucrose into hexoses.

Influence of 1-methylcyclopropene on pectic enzyme activities, gene expression, and cell wall modification in papaya (Carica papaya cv. ‘Sunrise’) fruit

SUMMARY ‘Sunrise’ papaya fruit harvested at two stages of maturity [colour break (< 10% yellow peel colour) and 25% yellow peel colour] were treated with 100 nl l–1 1-methylcyclopropene (1-MCP) to determine its effects on ripening, on the activities and levels of gene expression of polygalacturonase (PG), pectin methyl esterase (PME), and βgalactosidase ( βGal), and on the degradation of cell wall components. 1-MCP delayed ripening and the onset of the climacteric, although the peak in the respiration rate was almost the same as that in untreated control fruit. Colour-break fruit treated with 1-MCP exhibited a continuous increase in ethylene production, but at a lower rate than in control fruit. Consequently, 1-MCP-treated fruit ripened with a concomitant reduction in firmness, which was accompanied by an increase in PG and βGal enzyme activities and gene expression. On the other hand, fruit treated with 1-MCP at the 25% yellow stage exhibited lower levels of ethylene production and developed pulp with a rubbery texture at the ripe stage which was attributed to reduced PG, βGal, and PME enzyme activities and gene expression. This was consistent with the higher level of cell wall polysaccharides measured in 1-MCP-treated fruit. The above results indicated that ‘Sunrise’ papaya fruit can be treated with 1-MCP at the colour break stage since they have a greater capacity to recover from the effects of 1-MCP than fruit treated at the 25% yellow stage.

Metabolomics analysis of 'Housui' Japanese pear flower buds during endodormancy reveals metabolic suppression by thermal fluctuation

Dormancy is a complex phenomenon that allows plants to survive the winter season. Studies of dormancy have recently attracted more attention due to the expansion of temperate fruit production in areas under mild winters and due to climate changes. This study aimed to identify and characterize the metabolic changes induced by chilling temperatures, as well as during thermal fluctuation conditions that simulate mild winter and/or climate change scenarios. To do this, we compared the metabolic profile of Japanese pear flower buds exposed to constant chilling at 6 °C and thermal fluctuations of 6 °C/18 °C (150 h/150 h) during endodormancy. We detected 91 metabolites by gas chromatography paired with time-of-flight mass spectrometry (GC-TOF-MS) that could be classified into eight groups: amino acids, amino acid derivatives, organic acids, sugars and polyols, fatty acids and sterols, phenol lipids, phenylpropanoids, and other compounds. Metabolomics analysis revealed that the level of several amino acids decreased during endodormancy. Sugar and polyol levels increased during endodormancy during constant chilling and might be associated with chilling stress tolerance and providing an energy supply for resuming growth. In contrast, thermal fluctuations produced low levels of metabolites related to the pentose phosphate pathway, energy production, and tricarboxylic acid (TCA) cycle in flower buds, which may be associated with failed endodormancy release. This metabolic profile contributes to our understanding of the biological mechanism of dormancy during chilling accumulation and clarifies the metabolic changes during mild winters and future climate change scenarios.

Effects of Pre-chilling Potassium Nitrate and Post-chilling Hydrogen Cyanamide Application on Carbohydrate and Water Dynamics of ‘Housui’ Japanese Pear Spur Buds During Dormancy Under Mild Winter Conditions

Japanese pear ( Pyrus pyrifolia Nakai) trees were treated with potassium nitrate (KNO 3 ) before the onset of chilling and hydrogen cyanamide (CH 2 N 2 ) after the trees had accumulated 600 chilling hours (CH) to determine the effects on bud dormancy release, carbohydrate dynamics, and water content. For the carbohydrate and water content analyses, Japanese pear spur buds were collected after 0, 300, and 600 CH, and after the accumulation of 2000, 4000, 6000, and 8000 growing degree hours (GDH) during the dormancy period. Bud burst and flowering were hastened by both KNO 3 and CH 2 N 2 application, and both chemicals affected carbohydrate and water dynamics during dormancy. Sucrose concentration tended to increase during the chilling period, and to decrease under forcing conditions. Hexose concentrations increased in buds treated with KNO 3 , CH 2 N 2 , or both, and were associated with advances in bud burst. Sorbitol concentration decreased early in buds treated with CH 2 N 2 , suggesting rapid sorbitol catabolism. Water content gradually increased under forcing conditions in all buds, although those treated with both KNO 3 and CH 2 N 2 exhibited the highest values. Accordingly, we recommend the use of KNO 3 and CH 2 N 2 , before and after chilling, respectively, to promote the release of bud dormancy and increase bud burst rate in ‘Housui’ Japanese pear grown in regions with mild winter conditions.