Multiple non-specific effects of sphingosine on adenylate cyclase and cyclic AMP accumulation in S49 lymphoma cells preclude its use as a specific inhibitor of protein kinase C

John A Johnson, R. B. Clark

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Abstract

Recent studies with phorbol esters have suggested that protein kinase C (PKC) may play a role in the regulation of adenylate cyclase in mammalian cells. Since D-sphingosine has been reported to specifically inhibit PKC in many cell types, we evaluated its effects on stimulation of cyclic AMP accumulation by adrenaline in S49 lymphoma cells. We found sphingosine to have multiple non-specific effects which could not be explained by an inhibition of PKC. These effects included: (i) inhibition by sphingosine (50 μM) of adrenaline-stimulated cyclic AMP accumulation and sphingosine permeation of the cells which rendered them leaky to ATP; (ii) sphingosine (20 μM) augmentation of adrenaline-stimulated cyclic AMP accumulation; (iii) inhibition by sphingosine of adrenaline-stimulated adenylate cyclase in isolated membranes by up to 95%; and (iv) sphingosine (20 μM) inhibition of cellular mechanisms for the elimination of cyclic AMP. These results demonstrate the importance of evaluating the non-specific effects of sphingosine before concluding that its actions are the consequence of a specific inhibition of PKC.

Original languageEnglish (US)
Pages (from-to)507-511
Number of pages5
JournalBiochemical Journal
Volume268
Issue number2
DOIs
StatePublished - Jan 1 1990

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Sphingosine
Adenylyl Cyclases
Cyclic AMP
Protein Kinase C
Lymphoma
Epinephrine
Phorbol Esters
Permeation
Adenosine Triphosphate
Cells
Membranes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

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abstract = "Recent studies with phorbol esters have suggested that protein kinase C (PKC) may play a role in the regulation of adenylate cyclase in mammalian cells. Since D-sphingosine has been reported to specifically inhibit PKC in many cell types, we evaluated its effects on stimulation of cyclic AMP accumulation by adrenaline in S49 lymphoma cells. We found sphingosine to have multiple non-specific effects which could not be explained by an inhibition of PKC. These effects included: (i) inhibition by sphingosine (50 μM) of adrenaline-stimulated cyclic AMP accumulation and sphingosine permeation of the cells which rendered them leaky to ATP; (ii) sphingosine (20 μM) augmentation of adrenaline-stimulated cyclic AMP accumulation; (iii) inhibition by sphingosine of adrenaline-stimulated adenylate cyclase in isolated membranes by up to 95{\%}; and (iv) sphingosine (20 μM) inhibition of cellular mechanisms for the elimination of cyclic AMP. These results demonstrate the importance of evaluating the non-specific effects of sphingosine before concluding that its actions are the consequence of a specific inhibition of PKC.",
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AU - Clark, R. B.

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N2 - Recent studies with phorbol esters have suggested that protein kinase C (PKC) may play a role in the regulation of adenylate cyclase in mammalian cells. Since D-sphingosine has been reported to specifically inhibit PKC in many cell types, we evaluated its effects on stimulation of cyclic AMP accumulation by adrenaline in S49 lymphoma cells. We found sphingosine to have multiple non-specific effects which could not be explained by an inhibition of PKC. These effects included: (i) inhibition by sphingosine (50 μM) of adrenaline-stimulated cyclic AMP accumulation and sphingosine permeation of the cells which rendered them leaky to ATP; (ii) sphingosine (20 μM) augmentation of adrenaline-stimulated cyclic AMP accumulation; (iii) inhibition by sphingosine of adrenaline-stimulated adenylate cyclase in isolated membranes by up to 95%; and (iv) sphingosine (20 μM) inhibition of cellular mechanisms for the elimination of cyclic AMP. These results demonstrate the importance of evaluating the non-specific effects of sphingosine before concluding that its actions are the consequence of a specific inhibition of PKC.

AB - Recent studies with phorbol esters have suggested that protein kinase C (PKC) may play a role in the regulation of adenylate cyclase in mammalian cells. Since D-sphingosine has been reported to specifically inhibit PKC in many cell types, we evaluated its effects on stimulation of cyclic AMP accumulation by adrenaline in S49 lymphoma cells. We found sphingosine to have multiple non-specific effects which could not be explained by an inhibition of PKC. These effects included: (i) inhibition by sphingosine (50 μM) of adrenaline-stimulated cyclic AMP accumulation and sphingosine permeation of the cells which rendered them leaky to ATP; (ii) sphingosine (20 μM) augmentation of adrenaline-stimulated cyclic AMP accumulation; (iii) inhibition by sphingosine of adrenaline-stimulated adenylate cyclase in isolated membranes by up to 95%; and (iv) sphingosine (20 μM) inhibition of cellular mechanisms for the elimination of cyclic AMP. These results demonstrate the importance of evaluating the non-specific effects of sphingosine before concluding that its actions are the consequence of a specific inhibition of PKC.

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