Abstract
We have studied in vitro the effects of ethanol on the different enzymes involved in the biosynthesis of phosphatidylcholine (PC) via CDP-choline. Ethanol alters neither choline kinase (CK) nor CTP:phosphocholine cytidylyltransferase (CT) activities but, at levels higher than 50 mM, it does significantly inhibit microsomal cholinephosphotransferase (CPT) activity concomitantly with an increase in the ethanol concentration. A study of the kinetics of the reaction catalysed by CPT shows that ethanol decreases Vmax without altering Km, indicating a non-competitive inhibitory effect. An analysis of the thermodependence of CPT activity in the absence of ethanol reveals a break in the Arrhenius plot and thus a straight relationship between enzyme activity and the physico-chemical state of the microsomal membrane. Incubation of microsomes in the presence of ethanol increased the transition temperature from 25.8–28.2°C. Microsomes were also incubated with n-alkanols with chain-lengths of fewer than five carbon atoms at concentrations which, according to their partition coefficients, produce equimolar levels in the membrane. Under these conditions all the alkanols caused the same inhibitory effect. All these results demonstrate that ethanol modulate the PC biosynthesis at the level of CPT activity and does not affect the CT enzyme. The inhibition found on CPT is clearly dependent on the alteration produced by ethanol on the hepatic microsomal membrane.
Similar content being viewed by others
References
Lieber CS, Robins R, Li J, DeCarli LM, Mak KM, Fasulo JM, Leo MA: Phosphatidylcholine protects against fibrosis and cirrhosis in the baboon. Gastroenterology 106: 152–159, 1994
Carrasco MP, Sánchez-Amate MC, Segovia JL, Marco C: Studies on phospholipid biosynthesis in hepatocytes from alcoholic rats by using radiolabelled exogenous precursors. Lipids 31: 393–397, 1996
Lamb RG, Koch JC, Snyder JW, Huband SM, Bush SR: An in vitro model of ethanol-dependent liver cell injury. Hepatology 19: 174–182, 1994
Carrasco MP, Segovia JL, Marco C: Incorporation of exogenous precursors into neutral lipids and phospholipids in rat hepatocytes: Effect of ethanol in vitro. Biochem Pharmacol 56: 1639–1644, 1998
Carrasco MP, Sánchez-Amate MC, Marco C, Segovia JL: Evidence of differential effects produced by ethanol on specific phospholipid biosynthetic pathways in rat hepatocytes. Br J Pharmacol 119: 233–238, 1996
Chin JH, Goldstein DB: Drug tolerance in biomembranes: A spin label study of the effects of ethanol. Science 196: 684–685, 1977
Hitzemann RJ, Schueler HE, Graham-Brittain C, Kreishman GP: Ethanol-induced changes in neuronal membrane order. An NMR study. Biochim Biophys Acta 859: 189–197, 1986
Barry JA, Gawrisch K: Direct NMR evidence for ethanol binding to the lipid-water interface of phospholipid bilayers. Biochemistry 33: 8082–8088, 1994
Chiou JS, Kuo CC, Lin SH, Kamaya H, Ueda I: Interfacial dehydratation by alcohols: Hydrogen bonding of alcohols to phospholipids. Alcohol 8: 143–150, 1991
Covarrubias M, Rubin E: Ethanol selectively blocks a noninactivating K+ current expressed in Xenopus oocytes. Proc Natl Acad Sci USA 90: 6957–6960, 1993
Avdulov NA, Chochina SV, Daragan VA, Schroeder F, Mayo KH, Wood WG: Direct binding of ethanol to bovine serum albumin: A fluorescent and 13C NMR multiplet relaxation study. Biochemistry 35: 340–347, 1996
Pelech SL, Pritchard PH, Brindley DN, Vance DE: Fatty acids promote translocation of CPT:phosphocholine cytidylyltransferase to the endoplasmic reticulum and stimulate rat hepatic phosphatidylcholine synthesis. J Biol Chem 258: 6782–6788, 1983
Pelech SL, Power E, Vance DE: Activities of the phosphatidylcholine biosynthetic enzymes in rat liver during development. Can J Biochem Cell Biol 61: 1147–1152, 1983
Weinhold PA, Feldman DA: Choline-phosphate cytidylyltransferase. Meth Enzymol 209: 248–258, 1992
Vance DE, Pelech SD, Choy PC: CTP-phosphocholine cytidylyltransferase from rat liver. Meth Enzymol 71: 576–581, 1981
Cornell RB: Cholinephosphotransferase from mammalian sources. Meth Enzymol 209: 267–272, 1992
Lowry OH, Rosebrough NJ, Farr AL, Randall LJ: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–275, 1951
Morimoto K, Kanoh H: Acyl chain length dependency of diacylglycerol cholinephosphotransferase and diacylglycerol ethanolaminephosphotransferase. J Biol Chem 253: 5056–5060, 1978
Henneberry AL, McMaster CR: Cloning and expression of human choline/ethanolaminephosphotransferase: Synthesis of phosphatidylcholine and phosphatidylethanolamine. Biochem J 339: 291–298, 1999
Araki W, Wurtman R: Control of membrane phosphatidylcholine biosynthesis by diacylglycerol levels in neuronal cells undergoing neurite outgrowth. Proc Natl Acad Sci USA 94: 11946–11950, 1997
Coleman R, Bell RM: Phospholipid synthesis in isolated fat cells. J Biol Chem 252: 3050–3056, 1977
Tronchère H, Record M, Tercé F, Chap H: Phosphatidylcholine cycle and regulation on phosphatidylcholine biosynthesis by enzyme translocation. Biochim Biophys Acta 1212: 137–151, 1994
Sánchez-Amate MC, Carrasco MP, Zurera JM, Segovia JL, Marco C: Persistence of the effects of ethanol in vitro on the lipid order and enzyme activities of chick-liver membranes. Eur J Pharmacol 292: 215–221, 1995
Rothman A, Proverbio T, Proverbio F: Inhibitory effect of ethanol on the Na(+)-ATPase activity of rat kidney proximal tubular cell plasma membranes. Physiol Res 45: 205–211, 1996
Nieber K, Poelchen W, Sieler D, Illes P: Inhibition by ethanol of excitatory amino acid receptors in rat locus coeruleus neurons in vitro. Naunyn Schmiedebergs Arch Pharmacol 357: 299–308, 1998
Chu AJ: Mechanism by which ethanol inhibits phosphatidylcholine biosynthesis in human leukemic monocyte-like U937 cells. Cell Biochem Funct 12: 45–55, 1994
Weber FJ, Bont JAM: Adaptation mechanisms of microorganims to the toxic effects of organic solvents on membranes. Biochim Biophys Acta 1286: 225–245, 1996
Seeman P: The membrane actions of anesthetics and tranquilizers. Pharmacol Rev 24: 583–655, 1972
Sikkema J, de Bont JAM, Poolman B: Interactions of cyclic hydrocarbons with biological membranes. J Biol Chem 269: 8022–8028, 1994
Lyon RC, McComb JA, Schreurs J, Goldstein DB: A relationship between alcohol intoxication and the disordering of brain membranes by a series of short-chain alcohols. J Pharmacol Exp Ther 218: 669–675, 1981
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Carrasco, M.P., Segovia, J.L. & Marco, C. Modulation of biosynthesis of phosphatidylcholine via CDP-choline in rat liver: Influence of ethanol on the microsomal cholinephosphotransferase activity. Mol Cell Biochem 217, 43–50 (2001). https://doi.org/10.1023/A:1007271915160
Issue Date:
DOI: https://doi.org/10.1023/A:1007271915160