MECHANISM BY WHICH ETHANOL INHIBITS PHOSPHATIDYLCHOLINE BIOSYNTHESIS IN HUMAN LEUKEMIC MONOCYTE-LIKE U937 CELLS

A previous study showing that ethanol (ETOH) blocked [H-3]choline inco rporation into phosphatidylcholine (PC) suggested an inhibition of PC biosynthesis in human leukemic monocyte-like U937 cells. The mechanism of the inhibitory action of ETOH was investigated. Cells were pulsed with [H-3]choline for 30 min and chased in the presence or absence of ETCH for up to 6 h. PC biosynthesis was inhibited drastically within 1 h after exposure to ETOH which increased intracellular cAMP appreciab ly. After a 3-h treatment, ETOH significantly inhibited both choline k inase (CK) and the cytosolic CTP: cholinephosphate cytidylyltransferas e (CT). The inactivated CT was no longer stimulated by exogenous phosp hatidylglycerol (PG). There was no evidence for redistribution of CT a ctivity between cytosol and microsomes. When cells were exposed to 8-B romo-cAMP ranging from 100 to 300 mu M, PC biosynthesis remained unaff ected despite the drastically elevated cAMP. These results seem to sug gest that the raised cAMP is not a prerequisite for the inhibition of PC biosynthesis in U937 cells. Following pretreatment with protein kin ase inhibitors (H-89 and K-252a), PC biosynthesis was decreased signif icantly and the inhibitory effect of ETOH was potentiated. Taken toget her, our results suggest that the inhibition of PC biosynthesis and th e inhibitory effect of ETOH are independent of the activation of cAMP- dependent protein kinase. Unlike protein kinase inhibitors, pretreatme nt with tyrosine kinase inhibitors (erbstatin, genistein and tyrphosti n 25) resulted in differential effects on PC biosynthesis and on the i nhibitory action of ETOH. Genistein stimulated PC biosynthesis by 30 p er cent as well as partially reventing /reversing the ETOH action, whi le tyrphostin 25 produced a synergistic inhibition. The relevance of t yrosine phosphorylation/dephosphorylation to the regulation of PC bios ynthesis and ETOH action remains to be established.