Acceleration of Membrane Senescence in Cut Carnation Flowers

The lipid microviscosity of microsomal membranes from senescing cut carnation (Dianthus caryophyllus L. cv. White Sim) flowers rises with advancing senescence. The increase in membrane microviscosity is initiated within 3 to 4 days of cutting the flowers and coincides temporaDly with petal-inroDling denoting the climacteric-like rise in ethylene production. Treatment of young cut flowers with aminoethoxyvinylglycine prevented the appearance of petal-inrolling and delayed the rise in membrane microviscosity until day 9 after cutting. When freshly cut flowers or aminoethoxyvinylglycine-treated flowers were exposed to exogenous ethylene (1 microliter per liter), the microviscosity of microsomal membranes rose sharply within 24 hours, and inrofDing of petals was clearly evident. Thus, treatment with ethylene accelerates membrane rigidification. Silver thiosulphate, a potent anti-ethylene agent, delayed the rise in microsomal membrane microviscosity even when the flowers were exposed to exogenous ethylene. Membrane rigidification in both naturally senescing and ethylene-treated flowers was accompanied by an increased sterol:phosphoUpid ratio reflecting the selective loss of membrane phospholipid that accompanies senescence. The results collectively indicate that the climacteric-like surge in ethylene production during senescence of carnation flowers facilitates physical changes in membrane lipids that presumably lead to loss of membrane function. Cut flowers held in water senesce within 2 to 14 d depending upon the species (10). Aging of the flower petals is accompanied by morphological, biochemical, and biophysical deterioration. In carnations, for example, the earliest morphological indication of advancing senescence is a striking inrolling of the petals, a phenomenon that has been termed sleepiness (24), and in Ipomoea flowers, there is also an inrolling of the corolla (12). Invaginations of the tonoplast, thought to be a manifestation of autophagic activity by the vacuole, have also been observed during the early stages of morning glory senescence (16). In addition, flower senescence entails enhanced respiration and autolysis of the cell cytoplasm (18). Increased activities of such enzymes as RNase, DNase, and cell wall polysaccharide hydrolases are paralleled by a corresponding drop in the macromolecular constituents of cells (18). Evidence for an increase in membrane permeability during senescence of several flower species (1 1, 19, 24) suggests that there ' Supported in part by the Natural Sciences and Engineering Research Council of Canada. 2 Permanent address: Department of Biology, University of Waterloo, Waterloo, Ontario, Canada. To whom reprint requests should be addressed. is disruption of membrane integrity and loss of intracellular compartmentation. Kende et al. (3, 28) have noted a strong correlation between membrane leakiness and phospholipid breakdown in senescing flowers. Moreover, treatment of Tradescantia with ethylene accelerates the onset of membrane leakiness and phospholipid deterioration in petals, but the ethylene effect is dependent upon synthesis of new protein (28). Borochov et al. (6, 7) have reported that the microviscosity of plasma membranes from rose petals rises with advancing senescence in a manner that correlates with an increase in sterol:phospholipid ratio reflecting phospholipid breakdown. Senescing carnation flowers exhibit a climacteric-like rise in ethylene production (5, 17, 20). In addition, exposure of carnation flowers to exogenous ethylene induces inrolling of the petals and results in increased ethylene synthesis (10, 24, 26). In the present study, we have used cut carnation flowers to examine the ability of ethylene to induce chemical and physical changes in microsomal membrane lipids of senescing petals. MATERIALS AND METHODS Plant Material. Carnation flowers (Dianthus caryophyllus L. cv. White Sim) were grown in raised beds in a greenhouse according to established culturing procedures. Mature flowers were cut at the commercial stage of development (fully open with a yellowish tinted center) and either used directly for membrane isolation or trimmed to an 8 cm stem length and placed individually in 20-ml vials containing either deionized water or test solutions. Flowers held in water or test solutions were maintained at 22°C, and the levels of water or test solutions were adjusted as necessary to 1 cm below the calyx. Treatments. For treatment with ethylene, flowers were placed in deionized H20 in specially constructed Plexiglas chambers (135 L capacity) equipped with an internal fan to promote circulation, two ports for gas flow, and a removeable front panel. Exposure to ethylene was achieved by injecting ethylene into the chambers to a final concentration of 1 ,pl/l. Throughout the exposure, the chambers were connected to an air stream containing 1 ,ld/l ethylene that was flowing at 20 ml/min. Chambers containing control flowers were flushed at the same rate with air that had been rendered ethylene-free by passage through potassium permanganate coated with aluminum silicate (Purafil, Chamblee, GA). Ethylene treatments were terminated by removing the flowers from the chambers to a well-ventilated room. For measurements ofethylene, individual flowers were enclosed in round glass chambers (300 ml capacity) ventilated continuously with ethylene-free air (20 ml/min). Twice a day the air stream was disconnected, the inlet and outlet ports were plugged for I h, and a 4-ml sample was withdrawn from the chambers and introduced into a Varian gas chromatograph through a 2-ml sampling valve (17). The gas chromatograph was equipped with an alumina column (0.32 cm x 1.82 m) and operated at 40°C.