Sphingolipid metabolism and signal transduction: inhibition of in vitro phospholipase activity by sphingosine.
Sphingosine inhibits protein kinase C activity in vitro and has been used to implicate this enzyme in signal transduction and cell function. We report that sphingosine directly inhibits phospholipases A2 and D. Sphingosine inhibits Ca(2+)-dependent phospholipases A2 from Naja naja, porcine pancreas, Crotalus adamanteus, human disc and neutrophil in a dose-dependent manner with IC50 values ranging from 5-40 microM using [1-14C]oleate-labelled autoclaved E. coli (20 microM) as substrate. Inhibition is comparable using the same concentrations (20 microM) of [1-14C]oleate-labelled C. albicans or E. coli, or aqueous dispersions of 1-acyl-2-[1-14C]linoleoylglycerophosphoethanolamine or -choline. Sphinganine and stearylamine are as inhibitory as sphingosine; monoolein is less inhibitory (IC50 = 70 microM), while octylamine, N-acetylsphingosine, sphingomyelin and ceramide have no effect. Inhibition is relieved by increasing concentrations of substrate phospholipid. The molar ratio of sphingosine to phospholipid required for 50% inhibition ranges from 0.5 to 1.0 with 2-100 microM E. coli phospholipid. In contrast, sphingosine has a biphasic effect on the hydrolysis of E. coli by S. chromofuscus phospholipase D; concentrations less than or equal to 25 microM stimulate activity while concentrations greater than 25 microM are inhibitory. Addition of Triton X-100 eliminates both the stimulatory and inhibitory effects of sphingosine on phospholipase D activity.
Franson
R C
RC
Department of Biochemistry, Virginia Commonwealth University, Richmond.
Harris
L K
LK
Ghosh
S S
SS
Rosenthal
M D
MD
D000818:Animals; D001667:Binding, Competitive; D006801:Humans; D017830:Octoxynol; D010739:Phospholipase D / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010741:Phospholipases A / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011092:Polyethylene Glycols; D015398:Signal Transduction; D013107:Sphingolipids / Q000378:metabolism*; D013110:Sphingosine / Q000378:metabolism / Q000494:pharmacology*; D013379:Substrate Specificity
D013107:Sphingolipids; D011092:Polyethylene Glycols; D017830:Octoxynol; D010741:Phospholipases A; D010739:Phospholipase D; D013110:Sphingosine