Involvement of Calcium and Calmodulin in Membrane Deterioration during Senescence of Pea Foliage

Abstract

ABSTRACrThe prospect that Ca+2 promotes senescence by activating al has been e d using cut pea (Pisan satiwm co Aska) foliasge a model system. Senescence was induced by severing 17-day-old plats from their roots and mainting them in aqueous test solu in the dark for an additiona 4 days. Treatment of the foliage with the Ca2+ ionophore (A23187) during the senescence-inductio period promoted a laterd phase separtion of the bulk lipids in mirosom membranes indicating that intention of Ca2+ failitates membrae deterioation. In addition, microsomal membres from ionophore-treated tissue displayed an in capacity to convert 1-aminocyclopropane-l-carboxylic acid to ethylene and an increased propensity to produce the superoxide anion (02-). Treatment (23). These compounds are able to translocate Ca2+ from an aqueous phase across the intact bilayer into another aqueous compartment, are permselective with respect to the two major divalent cations of physiological fluids (Ca > Mg), and do not lyse the membrane. This observation is of particular significance in the context of senescing tissues because there is now evidence that free radical-mediated peroxidation of membrane lipids is a characteristic feature of senescence (3). Moreover, products of membrane lipid peroxidation, which would include oxidized di-and trienoic fatty acids, apparently accumulate in membrane matrices with advancing senescence and contribute to lateral phase separations in the lipid bilayers of senescing membranes (1).Exogenous Ca2' has been shown to delay senescence of apple tissue slices, in particular ethylene production and the onset of lipid peroxidation (9). However, this effect has been attributed to the ability of Ca2" to rigidify the surfaces of lipid bilayers by acting as a divalent ligand (7)

An ethylene-induced cDNA encoding a lipase expressed at the onset of senescence

“…This has been demonstrated by fluorescence depolarization and electron spin resonance for microsomal membranes from senescing cotyledons, flowers, leaves, and ripening fruit (5)(6)(7)(8) and for plasmalemma of ripening fruit and senescing flowers (5). The decrease in lipid fluidity is engendered by an enrichment of free sterols relative to fatty acids in the bilayer as fatty acids are cleaved from the membrane lipids and selectively removed, reflecting the fact that free sterols are known to restrict the mobility of phospholipid fatty acids (9).…”

PHOSPHOLIPID CATABOLISM AND SENESCENCE OF PEA FOLIAGE MEMBRANES: PARAMETERS OF Ca2+:CALMODULIN:PHOSPHOLIPASE A2−INDUCED CHANGES

“…In one experimental series, plants were treated with a 10~^ M CaCl2 solution containing 50/^M calcium ionophore (A 23187) prepared in a small volume of solvent (Leshem et al, 1984b). During the growth period, plants were sprayed with this solution every 3 d and, after 17 d, the severed foliage was maintained in darkness at 20 °C for an additional 4 d in beakers containing the Ca-ionophore.…”

“…^ • i i Measurements of ethylene production from its 1-ammocyclopropane-lcarboxylic acid precursor by isolated membranes were carried out essentially as described previously (Leshem et al, 1984b) Reaction -^^^^^ ^^^;;;^7^;;m embrane, buffer and the precursor are detailed m the legends to Tables and Figures The calmodulin inhibitor here employed was fiuphenazme. When assessing effects of stearic and linolenic acids, an ^^^^-^^^"^^"^5^!^^°" ^^[[f. '.…”

“…Microsomal membranes were isolated from 20 g pea leaves essentially as described previously (Leshem et al, 1984b). The final washed membrane preparation was then dialyzed overnight at 6 °C against four changes of 2 mM N-2-hydroxyethylpiperazine propane sulphonic acid (Epps) buffer, pH 8 5, in order to remove an ethylene inhibitor.…”

“…Onset of senescence could be attributed to a Ca^'^'-triggered activation of phospholipase A2 by formation of a Ca^"^: calmodulin: phospholipase A^ complex which, as detected by wide angle X-ray diffraction, causes a transition of bulk bilamellar membranal phospholipids from a liquid crystalline to a solid gel state (Leshem, 1984;Leshem, Sridhara & Thompson, 1984b). Concerning membranal sterols, it is assumed that their orientation is such that their hydroxyl groups, usually at the 3-position, are inserted in the region of the polar phospholipid heads while the planar cyclopentano-perhydrophenanthrene skeleton penetrates the lipid bilayer and is positioned and parallel to the fatty acid chains.…”