Modulation of human type II secretory phospholipase A2 by sphingomyelin and annexin VI

Abstract

Conjectural results have been reported on the capacity of inflammatory secreted phospholipase A2 (sPLA2) to hydrolyse mammalian membrane phospholipids. Development of an assay based on the release of non-esterified fatty acids by the enzyme acting on the organized phospholipid mixture constituting the membrane matrix has led to the identification of two prominent effectors, sphingomyelin (SPH) and annexin. Recombinant human type II sPLA2 hydrolyses red-cell membrane phospholipids with a marked preference for the inner leaflet. This preference is apparently related to the high content of SPH in the outer leaflet, which inhibits sPLA2. This inhibition by SPH is specific for sPLA2. Cholesterol counteracts the inhibition of sPLA2 by SPH, suggesting that the SPH-to-cholesterol ratio accounts in vivo for the variable susceptibility of cell membranes to sPLA2. Different effects were observed of the presence of the non-hydrolysable D-alpha-dipalmitoyl phosphatidylcholine (D-DPPC), which renders the membranes rigid but does not inhibit sPLA2. Annexin VI was shown, along with other annexins, to inhibit sPLA2 activity by sequestering the phospholipid substrate. The present study has provided the first evidence that annexin VI, in concentrations that inhibit hydrolysis of purified phospholipid substrates, stimulated the hydrolysis of membrane phospholipids by sPLA2. The activation requires the presence of membrane proteins. The effect is specific for type II sPLA2 and is not reproducible with type I PLA2. The activation by annexin VI of sPLA2 acting on red cell membranes results in the preferential release of polyunsaturated fatty acids. It suggests that type II sPLA2, in conjunction with annexin VI, might be involved in the final step of endocytosis and/or exocytosis providing the free polyunsaturated fatty acids acting synergistically to cause membrane fusion.

Modulation of the activity of cytosolic phospholipase A2α (cPLA2α) by cellular sphingolipids and inhibition of cPLA2α by sphingomyelin

“…Subbaiah and Liu ( 25 ) reported that lecithin-cholesterol acyltransferase, a specialized phospholipase A responsible for the esterifi cation of cholesterol in plasma, was inhibited by SM. The activities of lipoprotein lipase and secretory PLA2 have also been shown to be inhibited by SM in vitro (26)(27)(28)(29)(30). Dawson et al ( 31 ) reported that diacylglycerol-stimulated intracellular phospholipases were strongly inhibited by SM.…”

The Functions of Five Distinct Mammalian Phospholipase A2s in Regulating Arachidonic Acid Release

“…Proinflammatory stimuli may cause alterations in membrane lipid packing and asymmetry, which are thought to induce increased cellular sPLA 2 -IIA sensitivity. More recently, sphingomyelin turnover, one of the primary responses following ligation of the receptors for proinflammatory cytokines (67,68), has been proposed to perturb the outer leaflet of the membrane, thereby modulating the activity of sPLA 2 s acting on biological membranes (69,70).…”

Mechanisms by Which Elevated Intracellular Calcium Induces S49 Cell Membranes to Become Susceptible to the Action of Secretory Phospholipase A2

“…Ref. [52][53][54][55]. One possible resolution to that controversy is that specificity is determined not by the structure of the head group per se, but by the physical state of the phospholipids within the bilayer and that an altered distribution of phospholipids across the bilayer could affect that physical state (52).…”

“…Indeed, some cells appear to be more readily hydrolyzed by sPLA 2 following pre-hydrolysis of the plasma membrane with sphingomyelinase (27,55,61). We examined this hypothesis briefly by incubating S49 cells with 1 unit/ml sphingomyelinase for various times at 37°C and then assaying the time course of membrane hydrolysis in the presence of sPLA 2 .…”