The role of protein kinase C in cellular tolerance to ethanol.
We have shown that ethanol inhibits uptake of adenosine by a specific nucleoside transporter in NG108-15 neuroblastoma x glioma cells and that cAMP-dependent protein kinase (PKA) activity is required for this inhibition. After chronic exposure to ethanol, adenosine uptake is no longer inhibited on rechallenge with ethanol, i.e. transport has become tolerant to ethanol. Here we show that protein kinase C (PKC) contributes to ethanol-induced tolerance of adenosine transport. Activation of PKC by phorbol esters in control cells results in an ethanol-tolerant phenotype, similar to that produced by chronic ethanol exposure. In addition, chronic exposure to ethanol increases the amounts of alpha, delta, and epsilon PKC. However, reducing PKC activity by inhibition with chelerythrine during chronic exposure to ethanol or down-regulation by phorbol esters prevents the development of ethanol-induced tolerance of adenosine transport. By contrast, the inhibition of PKA activity produces tolerance to ethanol inhibition of adenosine uptake. When protein phosphatase inhibitors are present, inhibiting PKA activity has no effect on ethanol sensitivity of adenosine uptake, suggesting a role for protein phosphatases in the regulation of ethanol sensitivity of uptake. Taken together, our results suggest that PKA and PKC have opposing effects on the ethanol sensitivity of adenosine transport; PKA activity is required for ethanol sensitivity, and PKC activation produces tolerance. Based on these data, we propose that chronic ethanol exposure increases PKC activity, leading to the activation of a protein phosphatase (1 or 2A). This phosphatase then dephosphorylates a PKA-phosphorylated site, which is required for ethanol to inhibit adenosine uptake. Therefore, the sensitivity of adenosine transport to ethanol appears to be maintained by a balance of PKA and protein phosphatase activities, and PKC may regulate phosphatase activity.
Coe
I R
IR
Ernest Gallo Clinic and Research Center, University of California, San Francisco, California 94110, USA.
Yao
L
L
Diamond
I
I
Gordon
A S
AS
D000241:Adenosine / Q000378:metabolism; D001692:Biological Transport; D015536:Down-Regulation; D004361:Drug Tolerance; D004789:Enzyme Activation; D000431:Ethanol / Q000494:pharmacology*; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D000431:Ethanol; D011493:Protein Kinase C; D000241:Adenosine