Sphingolipids and cholesterol modulate membrane susceptibility to cytosolic phospholipase A(2).
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-beta-cyclodextrin (5 mm) 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 vivo experimental approaches, indicate that membrane microdomains enriched in SPH and Chol play a role in the modulation of the activity of cPLA2 and in arachidonic acid-derived mediator production.
Klapisz
E
E
Laboratoire de Biochimie, UPRES-A 7079, Université Pierre et Marie Curie, CHU Saint Antoine, F-75571 Paris, Cedex 12, France.
Masliah
J
J
Béréziat
G
G
Wolf
C
C
Koumanov
K S
KS
D000818:Animals; D016466:CHO Cells; D000001:Calcimycin; D002518:Ceramides / Q000494:pharmacology; D002784:Cholesterol / Q000494:pharmacology*; D006224:Cricetinae; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004837:Epinephrine / Q000494:pharmacology; D005227:Fatty Acids / Q000032:analysis / Q000378:metabolism; D006801:Humans; D007476:Ionophores / Q000494:pharmacology; D008051:Lipid Bilayers / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism*; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D051381:Rats; D018341:Receptors, Adrenergic, alpha-2; D011994:Recombinant Proteins; D013107:Sphingolipids / Q000494:pharmacology*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000494:pharmacology
C547771:ADRA2B protein, human; D002518:Ceramides; D004791:Enzyme Inhibitors; D005227:Fatty Acids; D007476:Ionophores; D008051:Lipid Bilayers; D008081:Liposomes; D008563:Membrane Lipids; C064769:N-acetylsphingosine; D010743:Phospholipids; D018341:Receptors, Adrenergic, alpha-2; D011994:Recombinant Proteins; D013107:Sphingolipids; C101954:N-caproylsphingosine; D000001:Calcimycin; D002784:Cholesterol; D010741:Phospholipases A; D013110:Sphingosine; D004837:Epinephrine