Cysteine protease gene expression and proteolytic activity during senescence of Alstroemeria petals

The functional life of the flower is terminated by senescence and/or abscission. Multiple processes contribute to produce the visible signs of petal wilting and inrolling that typify senescence, but one of the most important is that of protein degradation and remobilization. This is mediated in many species through protein ubiquitination and the action of specific protease enzymes. This paper reports the changes in protein and protease activity during development and senescence of Alstroemeria flowers, a Liliaceous species that shows very little sensitivity to ethylene during senescence and which shows perianth abscission 8-10 d after flower opening. Partial cDNAs of ubiquitin (ALSUQ1) and a putative cysteine protease (ALSCYP1) were cloned from Alstroemeria using degenerate PCR primers and the expression pattern of these genes was determined semi-quantitatively by RT-PCR. While the levels of ALSUQ1 only fluctuated slightly during floral development and senescence, there was a dramatic increase in the expression of ALSCYP1 indicating that this gene may encode an important enzyme for the proteolytic process in this species. Three papain class cysteine protease enzymes showing different patterns of activity during flower development were identified on zymograms, one of which showed a similar expression pattern to the cysteine protease cDNA.

Programmed cell death (PCD) processes begin extremely early in Alstroemeria petal senescence

-In the Liliaceous species Alstroemeria, petal senescence is characterized by wilting and inrolling, terminating in abscission 8-10 d after flower opening. -In many species, flower development and senescence involves programmed cell death (PCD). PCD in Alstroemeria petals was investigated by light (LM) and transmission electron microscopy (TEM) (to study nuclear degradation and cellular integrity), DNA laddering and the expression programme of the DAD-1 gene. -TEM showed nuclear and cellular degradation commenced before the flowers were fully open and that epidermal cells remained intact whilst the mesophyll cells degenerated completely. DNA laddering increased throughout petal development. Expression of the ALSDAD-1 partial cDNA was shown to be downregulated after flower opening. -We conclude that some PCD processes are started extremely early and proceed throughout flower opening and senescence, whereas others occur more rapidly between stages 4-6 (i.e. postanthesis). The spatial distribution of PCD across the petals is discussed. Several molecular and physiological markers of PCD are present during Alstroemeria petal senescence. © New Phytologist (2003).

Characterization of a novel lipoxygenase-independent senescence mechanism in Alstroemeria peruviana floral tissue

The role of lipoxygenase (lox) in senescence ofAlstroemeria peruviana flowers was investigated using a combination of in vitro assays and chemical profiling of the lipid oxidation products generated. Phospholipids and galactolipids were extensively degraded during senescence in both sepals and petals and the ratio of saturated/unsaturated fatty acids increased. Lox protein levels and enzymatic activity declined markedly after flower opening. Stereochemical analysis of lox products showed that 13-lox was the major activity present in both floral tissues and high levels of 13-keto fatty acids were also synthesized. Lipid hydroperoxides accumulated in sepals, but not in petals, and sepals also had a higher chlorophyll to carotenoid ratio that favors photooxidation of lipids. Loss of membrane semipermeability was coincident for both tissue types and was chronologically separated from lox activity that had declined by over 80% at the onset of electrolyte leakage. Thus, loss of membrane function was not related to lox activity or accumulation of lipid hydroperoxides per se and differs in these respects from other ethylene-insensitive floral tissues representing a novel pattern of flower senescence.

Increasing flower longevity in Alstroemeria

The vase-life of Alstroemeria (cv. Rebecca) flowers is terminated when the tepals abscise. Abscission was accelerated by both chloroethylphosphonic acid (CEPA) and 1-aminocyclopropane-1-carboxylic acid (ACC). Petals abscised 24 h earlier compared with controls, when isolated cymes were placed in 340 nM CEPA, and earlier still when higher concentrations were used. This suggests that flowers of this Alstroemeria cultivar are very ethylene sensitive. Treatment with silver thiosulphate (STS) overcame the effects of exposure to CEPA and delayed perianth abscission of untreated isolated flowers by 3-4 days. The inclusion of 1% sucrose in the vase solution also extended longevity but not by as much as STS treatment; combined STS and sucrose treatments did not increase longevity beyond that of either treatment alone. However, removal of the young buds from the axil of the first flower was the most effective treatment to extend vase-life and encouraged the growth and development of the remaining flower. Flowers on cut inflorescences from which young axillary buds were trimmed more than doubled in fresh weight 6 days after flower opening compared with an increase of only 70-80% in those untreated or treated with STS and/or sucrose. Growth was less in isolated cymes but followed a similar pattern. The effect of STS and/or sucrose treatment was synergistic with the trimming treatment and thus the vase-life of trimmed, STS and sucrose-treated flowers was over 7 days longer than that for untreated controls. © 2003 Elsevier B.V. All rights reserved.