Erin M. O'Donoghue

Programmed cell death during flower senescence: isolation and characterization of cysteine proteinases from Sandersonia aurantiaca

Cysteine protease inhibitors delayed the senescence of Sandersonia aurantiaca Hook. flowers. Tepal fading and wilting occurred later in the 2,2´ -dipyridyl-treated flowers, and these flowers had a greater soluble protein content and less active endoproteases compared with control flowers that were held in water. Biochemical analysis revealed the presence of several protease-active bands in the soluble protein fraction of Sandersonia tepals. Activity of the polypeptides increased as flower senescence progressed. Western analysis with an antibody raised against the castor bean cysteine proteinase identified homologous proteins in Sandersonia flowers (ca 46, 41 and 31kDa). Three cDNAs encoding cysteine proteinases were isolated from Sandersonia tepals (PRT5, PRT15 and PRT22). Expression of all three increased in tepals as senescence progressed. mRNAs for PRT5 were detected only in senescing flower tissue, whereas PRT15 and PRT22 were expressed in leaf, stem and root tissue. PRT5 has significant homology to C-terminus KDEL proteins, which have a role in the degradation of plant cell contents during programmed cell death. PRT15 is most similar to cysteine proteinases with a long C-terminal extension, whereas PRT22 is homologous to stress-induced cysteine proteinases.

Physiological changes in asparagus spears immediately after harvest

Abstract Respiratory activity of asparagus spears measured in the field showed a 4-fold increase between the butt and the tip of 190-mm spears. After harvest, there was an immediate rise in the respiration rate, followed by a rapid drop during 24 h at 16°C to a constant level, ∼ 30% of the peak respiration rate. Strong gradients of sugars and proteins were measured along the spears with low levels of sugars and high levels of proteins present in the spear tips. Sugars declined markedly in the first 24 h after harvest, particularly in spear tips. Proteins in spear tips were unchanged 24 h after harvest, but had decreased 25% by 72 h. Total free amino acids remained steady for the first 24 h after harvest, but increased by 75% at 48 h. Asparagine/aspartic acid increased, whereas glutamine/glutamic acid and proline decreased in concentration substantially during the first 24 h after harvest. Tips of taller spears had a lower sugar content and more protein than tips of short spears.

Unravelling senescence : new opportunities for delaying the inevitable in harvested fruit and vegetables

Current technology for maintaining the postharvest quality of fresh produce relies on the precise control of storage temperature and/or atmosphere conditions. These technologies have evolved from over 70 years of largely empirical postharvest research. New knowledge about the mechanisms and control of senescence is emerging from biochemical and molecular studies, and is providing exciting opportunities to delay the inevitable loss of quality that accompanies the postharvest senescence of horticultural crops. This knowledge is generating elite plants that have altered postharvest characteristics, and will simplify the future development of effective postharvest technology.

Flavour and metabolic changes in asparagus during storage

Abstract Flavour acceptability of apical spear segments declined within 2 days of storage at 1°C and further declined after 7 days of storage. Decline in flavour acceptance was accompanied by off-flavour development, a decrease in soluble carbohydrates and an increase in protein content of spear tip segments. Respiration rate of spears stored at 1 or 20°C declined after harvest before reaching an equilibrium. A significant loss of soluble carbohydrates and proteins from tip segments occurred during storage at 20°C. Storage at 1°C reduced the rate of substrate loss.

Vase solutions containing sucrose result in changes to cell walls of sandersonia (Sandersonia aurantiaca) flowers

It is well established that vase solutions containing sugar can improve the vase-life of many cut flower crops. Since cut sandersonia flowers supplied with 2% sucrose are firmer during wilting compared to water-fed controls, we have examined whether the effects of sucrose treatment extend to alterations in cell wall structure in the floral tissues, which may influence the wilting-related flower softening. Mature but not fully opened individual flowers were removed from the stems of sandersonia plants and were fed continuously with either 2% sucrose solution or water for up to 10 days. Sucrose supplementation resulted in decreased amounts of chelator-soluble pectin and increased amounts of Na2CO3-soluble pectin per individual flower, and also changed the molecular size profiles of both these pectin fractions compared to the water-fed controls. The molecular size differences were obvious after 3 days in vase solutions, and diminished with subsequent vase time. Senescence-related galactose loss was delayed in sucrose-fed flowers but there was no difference in the levels of β-galactosidase activity present in these flowers compared to controls. The observed differences in cell wall pectins due to sucrose feeding were not reflected in differences to the overall firmness of pre-senescent flowers (up to day 3). High levels of galactose persisted into the wilting phase when sucrose-fed flowers were firmer than water-fed controls. We conclude that while sucrose induced significant quantitative and qualitative differences in pectin fractions and galactose content, firmness of floral tissue, particularly during senescence, was not governed by these events alone.

Characterization of the harvest-induced expression of β-galactosidase in Asparagus officinalis

Abstract The harvest-induced senescence of asparagus spears is accompanied by both up-regulated and down-regulated gene expression. The expression of pTIP31, coding for asparagus β -galactosidase (EC 3.2.1.23) is temporally associated with removal of the asparagus spear from the main body of the plant — neither wounding or compression treatments induce up-regulation of transcripts corresponding to pTIP31. Harvest-induced pTIP31 transcripts appear initially in cells below the meristems of the side branches, and in inner bracts and developing flowers of the spear tip, and in the cells of the outer cortex and conjunctive tissue lower down the spear. Transcripts are also located in vascular tissues 24 h after harvest. Enhanced levels of β -galactosidase activity are found in the branches and bracts of the spear tip within 12 h of harvest, while the rate of increase in activity in the middle zone of the spear was comparable to that of the branches and bracts after 24 h. The proportion of galactose in spear cell walls decreases after harvest. We propose that galactosidase activity releases galactose residues that may be used as respiratory substrate for the rapidly deteriorating asparagus spears.

Galactosidases in opening, senescing and water-stressed Sandersonia aurantiaca flowers

Three glycosyl hydrolase family 35 β-galactosidase-encoding cDNAs, SaGAL1 (full-length), SaGAL2 and SaGA3L (both partial), have been isolated from Sandersonia aurantiaca (Hook.) SaGAL1 protein was functionally expressed in E. coli and β-galactosidase identity confirmed by activity assay. All three clones are primarily expressed in tepal tissues of senescing sandersonia flowers. In order to identify relationships between tepal texture and galactose metabolism, cut sandersonia flowers were treated with sucrose, periods of dryness or PEG and parameters associated with galactose metabolism and firmness were monitored. Sucrose supplementation, known to increase tepal firmness, delayed expression of SaGAL1 and SaGAL3 in opening (stage 5) flowers, whereas the response to periods of dryness followed by rehydration depended on the maturity of the flower. These treatments also tended to hasten the onset of processes associated with programmed cell death, monitored by PRT5 (a senescence-associated protease) expression. Galactosidase activity and cell wall galactose content were also affected but in an inconsistent manner. PEG supplied to opening flowers for 1 d followed by water, induced a long period of wilt, and intensive PRT5 expression. However, β-galactosidase gene expression and activity was delayed in these flowers, and cell-wall galactose content changed apparently independently of galactosidase activity. We have not been able to demonstrate a causal connection between the change in petal texture and concurrent induction of galactose mobilisation in sandersonia during normal development and senescence.

Xyloglucan endotransglycosylase: a role after growth cessation in harvested asparagus

Little is known about the mode of xyloglucan endotransglycosylase (XET) activity in cell walls once the turgor, which drives expansion, is reduced. Such a situation exists when growing shoots are excised from the parent plant, and is the case for many commercially valuable vegetable crops, e.g. asparagus, Asparagus officinalis L. XET activity was present in all zones of rapidly growing, immature asparagus spears, but with highest levels at the spear base where elongation growth had ceased. Activity increased in all parts of the spear for up to 72 h after harvest. Two members of the XET-related gene family in asparagus (AoXET1 and AoXET2) were isolated and mRNA corresponding to these clones accumulated at low levels, particularly in the basal zone during spear growth. Transcript levels increased in all parts of the asparagus spear after harvest, but this increase did not coincide with the increase in XET activity. The harvest-related changes to xyloglucan molecular weight were restricted to slight, segment-specific, up- or down-shifts. However, this may hide strategic alterations to linkages leading to a more rigid wall without major changes in overall molecular weight. The initial postharvest surge in XET activity could be related to harvest stresses such as water deficit, but we propose that the later induction of AoXET1 and AoXET2 is linked to the development of lignified secondary cell walls.