Sphingolipids and cholesterol modulate membrane susceptibility to cytosolic phospholipase A2

Abstract

Modulation of cytosolic phospholipase A 2 (cPLA 2 ) activity by sphingomyelin (SPH), ceramide (Cer), and cholesterol (Chol) was investigated in CHO-2B cells activated by the calcium ionophore A23187 and epinephrine. Chol depletion of CHO-2B cells by treatment with methyl-␤cyclodextrin (5 m M ) resulted in the inhibition of the release of arachidonic acid whereas the restoration of the level by Chol-loaded cyclodextrin relieved inhibition. Conversion of CHO-2B cellular SPH to Cer by Staphylococcus aureus sphingomyelinase enhanced endogenous cPLA 2 activation as well as uptake by cells of C2-and C6-ceramide analogs. These results were confirmed in vitro with purified human recombinant cPLA 2 acting on a model phospholipid substrate. The enzyme activity was inhibited by SPH but reactivated by Cer as well as by Chol added to glycerophospholipid liposomal substrates containing SPH. The results of this study, which combine in situ and in vitro experimental approaches, indicate that membrane microdomains enriched in SPH and Chol play a role in the modulation of the activity of cPLA 2 and in arachidonic acid-derived mediator production. -Klapisz, E., J. Masliah, G. Béréziat, C. Wolf, and K. S. Koumanov. Sphingolipids and cholesterol modulate membrane susceptibility to cytosolic phospholipase A 2 .

Phospholipase A2-Induced Remodeling Processes on Liquid-Ordered/Liquid-Disordered Membranes Containing Docosahexaenoic or Oleic Acid: A Comparison Study

“…It is in accordance with our previous studies where SM was shown to decrease PLA 2 activity in different substrate-containing vesicles. 26,33 The activity of the enzyme (human recombinant cytosolic PLA 2 and human type II secretory PLA 2 ) was shown to be reduced in PC/SM containing mixtures compared to pure PC vesicles. In the present study, PLA 2 activity was examined as a function of the degree of fatty acid unsaturation.…”

“…Visualization of the coexisting phases was made possible by the use of the fluorescent lipid analogue egg Rhodamine-PE that preferentially partitions into L d phase. 33 When two liquid-disordered (L d ) and liquid-ordered (L o ) phases coexist in the plane of the membrane bilayer, the bright background is assigned to the L d phase, while the observed dark circular domains are assigned to the L o phase.…”

Regulation of the Activity and Fatty Acid Specificity of Lecithin-Cholesterol Acyltransferase by Sphingomyelin and Its Metabolites, Ceramide and Ceramide Phosphate

“…A recent report by Lee et al (10) showed that HDL from patients with Niemann-Pick disease, which contains a high concentration of SM, also exhibits impaired cholesterol esterification by LCAT and that this is mostly attributable to the lack of secretory SMase in these patients. However, studies with various phospholipases showed that ceramide, the product of SMase C reaction, is an independent activator of the phospholipases (12)(13)(14)(15)(16). Since LCAT is essentially a specialized phospholipase A that utilizes cholesterol as an acyl acceptor in place of water, this raised the possibility that the stimulation of LCAT reaction by SMase C treatment of lipoproteins is due to the formation of an activator (ceramide), rather than depletion of the inhibitor (SM).…”

“…Although the cellular functions of ceramide have been investigated well, its possible effects in the plasma compartment are unknown. Studies from several laboratories, including ours, have shown that ceramide is an activator of secretory and cytosolic phospholipases in vitro ( − ). Because of its tendency to form inverted hexagonal phase structures (H II ), it disrupts the membrane bilayer ( , ), and this may be the basis for its activation of phospholipase activities.…”