Molecular Species Specificity of Phospholipid Breakdown in Microsomal Membranes of Senescing Carnation Flowers

Abstract

ABSTRACrDuring senescence of cut canation flowers, there is extensive breakdown of microsomal phospholipid. This is attributable, at least in part, to lipolytic activity associated directly with the microsomal membranes. Evidence indicating that one or more of the lipid-degrading enzymes in these membranes preferentially degrade phospholipid molecular species containing two diunsaturated acyl chais or at least one polyunsaturated acyl chain has been obtained by using radiolabeled phosphatidylcholine the lipids more susceptible to attack by lipases.In the present study, we describe evidence indicating that the phospholipid-degrading enzymes associated with microsomal membranes of senescing carnation flowers exhibit specificity for phospholipid molecular species containing two diunsaturated fatty acids or at least one polyunsaturated fatty acid. This finding is discussed in the context that provision of specific molecular species of phospholipids, which serve as preferred substrates for membrane-associated lipases, may regulate the progression and rate of phospholipid degradation in senescing membranes. MATERIALS AND METHODSPlant Material and Membrane Isolation. Carnation flowers (Dianthus caryophyllus L. cv White Sim) were grown in a commercial greenhouse (Unsworth and Sons, Burlington, Ontario). They were cut at a young stage when the petals had expanded approximately 2 cm beyond the sepals. The stems were trimmed to a length of 22 cm, and the flowers were placed individually in glass culture tubes containing deionized water. They were maintained at 22°C under continuous illumination (240 ft-c) until they had reached specific stages of senescence, viz., stage II, flowers that still possessed yellowish-tinted centers, but were fully expanded; stage III, flowers that were completely white in color but were not yet showing petal-inrolling; stage IV, senescent flowers showing petal-inrolling.Microsomal membranes were isolated from stage II, III, and IV flowers in 10 mm Epps3 (pH 8.5), as described previously (25) and washed once by resuspension in the same buffer and centrifugation at 131,000g for 1 h. The resulting pellet of membranes was resuspended in 3 ml 70 mM Epps (pH 8.5), and dialyzed at 4°C against 3 changes of 600 ml of 2 mm Epps (pH 8.5), for 15 h. After dialysis, the protein concentration was adjusted to 1 mg ml-' with 70 mM Epps (pH 8.5). Protein was measured as described by Bradford (5).Phospholipid Degradation. The capability of isolated microsomal membranes to degrade various molecular species of phospholipids was determined by using radiolabeled substrates. An aliquot (1.8 ml) of washed, dialyzed membrane suspension (I mg protein ml-') was vortexed for 3 min with 0.

Decreased Membrane Integrity in Aging Typha latifolia L.Pollen (Accumulation of Lysolipids and Free Fatty Acids)

“…In the resulting LPC, the mo1 % of 16:O was considerably increased and that of 18:2 decreased, as compared with the mo1 % of these two fatty acids in the original PC ( Table 111). Assuming that the fatty acids 16:0,18:0, and 18:l are at the sn-1 position and that 18:3 is entirely at the sn-2 position ( e g Brown et al, 1987), the theoretical composition of sn-1-LPC is very similar to that found after phospholipase A2 incubation. This confirms the preference of 16:O in pollen PC for the sn-1 position.…”

Lipid Profiling Demonstrates That Suppressing Arabidopsis Phospholipase Dδ Retards ABA-Promoted Leaf Senescence by Attenuating Lipid Degradation

“…The disruption of membranes results primarily from the remarkable metabolism of membrane lipids. Loss of membrane phospholipids was observed during natural and ethylene-induced senescence of cut carnation flowers [31], [32]. Changes in the metabolic relationships among phospholipids, and between galactolipids and phospholipids, have been reported to occur during senescence-associated lipid breakdown.…”

Is Petal Senescence Due to Sugar Starvation?

“…Sugars or cycloheximide often cannot postpone senescence if they are applied after a fixed period from the onset of flower opening. Figure 2 shows the time line of a number of processes, both during starvation and during petal senescence (Wiemken et al, 1976;Brown et al, 1987;Stephenson and Rubinstein, 1998).…”