Phosphatidylethanolamine N-methyltransferase (PEMT) catalyzes the conversion of phosphatidylethanolamine to phosphatidylcholine (PC). We investigated whether there was diminished secretion of lipoproteins from hepatocytes derived from mice that lacked PEMT (Pemt ؊/؊ ) compared with Pemt ؉/؉ mice. Hepatocytes were incubated with 0.75 mM oleate, the media were harvested, and triacylglycerol (TG), PC, apolipoprotein (apo) B100, and apoB48 were isolated and quantified. Compared with hepatocytes from Pemt ؉/؉ mice, hepatocytes from Pemt ؊/؊ mice secreted 50% less TG, whereas secretion of PC was unaffected. Fractionation of the secreted lipoproteins on density gradients demonstrated that the decrease in TG was in the very low density lipoprotein (VLDL)/low density lipoprotein fractions. The secretion of apoB100 was decreased by ϳ70% in VLDLs/ low density lipoproteins, whereas there was no significant decrease in apoB48 secretion in any fraction. Transfection of McArdle hepatoma cells (that lack PEMT) with PEMT cDNA enhanced secretion of TG in the VLDLs. Because the levels of PC in the hepatocytes and hepatoma cells were unaffected by the lack of PEMT expression, there appears to be an unexpected requirement for PEMT in the secretion of apoB100-containing VLDLs.
Phosphatidylethanolamine N-methyltransferase (PEMT)is involved in a secondary pathway for production of phosphatidylcholine (PC) in liver. We fed Pemt ؊/؊ mice a high fat/high cholesterol diet for 3 weeks to determine whether or not PC derived from PEMT is required for very low density lipoprotein secretion. Lipid analyses of plasma and liver indicated that male Pemt ؊/؊ mice accumulated triacylglycerols in their livers and were unable to secrete the same amount of triacylglycerols from the liver as did Pemt ؉/؉ mice. Plasma levels of triacylglycerol and both apolipoproteins B100 and B48 were significantly decreased only in male Pemt ؊/؊ mice. Experiments in which mice were injected with Triton WR1339 showed that, whereas hepatic apoB100 secretion was decreased in male Pemt ؊/؊ mice, the decrease in plasma apoB48 in male Pemt ؊/؊ mice was not due to reduced secretion. Moreover, female and, to a lesser extent, male Pemt ؊/؊ mice showed a striking 40% decrease in plasma PC and cholesterol in high density lipoproteins. These results suggest that, even though the content of hepatic PC was normal in PEMT-deficient mice, plasma lipoprotein levels were profoundly altered in a gender-specific manner. Phosphatidylcholine (PC)1 is synthesized by the Kennedy pathway (CDP-choline pathway), by methylation of phosphatidylethanolamine (PE), or by acylation of lyso-PC (LPC). The Kennedy pathway is the major route for phosphatidylcholine (PC) synthesis in all mammalian tissues and is dependent on the intake of choline (1). In the liver an additional pathway for PC production is catalyzed by phosphatidylethanolamine Nmethyltransferase (PEMT), which converts PE to PC via the transfer of three methyl groups from S-adenosylmethionine (2). The liver-specific expression of PEMT suggests that this enzyme might play a role in bile secretion and/or very low density lipoprotein (VLDL) secretion. PC is the primary phospholipid of all classes of lipoproteins in mammals (3) and is required for the secretion of VLDL; other phospholipids cannot substitute (4 -7). The Kennedy pathway is required for maintaining normal plasma VLDL levels, because rats fed a choline-deficient diet for 3 days had a 6-fold increase in hepatic triacylglycerol (TG) and decreased plasma TG (6).The role of PEMT-derived PC for lipoprotein secretion/metabolism has not been clearly established. Treatment of hepatocytes with 3-deazaadenosine, an inhibitor of methylation reactions that utilize S-adenosylmethionine, inhibited PEMT activity by more than 90% but did not decrease VLDL secretion (8). Thus, PEMT activity did not appear to be required for VLDL secretion. Subsequent experiments revealed that one pool of hepatic PC, which was produced by the methylation of PE derived from phosphatidylserine (PS) decarboxylation, was preferentially incorporated into VLDL secreted by hepatocytes (9). Moreover, secretion of this pool of PC labeled by this pathway was not inhibited by 3-deazaadenosine even though this inhibitor reduced the incorporation of labeled serine into PC of hepa...
Mild hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Homocysteine, a nonprotein amino acid, is formed from S-adenosylhomocysteine and partially secreted into plasma. A potential source for homocysteine is methylation of the lipid phosphatidylethanolamine to phosphatidylcholine by phosphatidylethanolamine N-methyltransferase in the liver. We show that mice that lack phosphatidylethanolamine N-methyltransferase have plasma levels of homocysteine that are ϳ50% of those in wild-type mice. Hepatocytes isolated from methyltransferase-deficient mice secrete ϳ50% less homocysteine. Rat hepatoma cells transfected with phosphatidylethanolamine N-methyltransferase secrete more homocysteine than wild-type cells. Thus, phosphatidylethanolamine N-methyltransferase is an important source of plasma homocysteine and a potential therapeutic target for hyperhomocysteinemia.Mild hyperhomocysteinemia is an independent risk factor for cardiovascular (1, 2) and atherosclerotic disease (3). Total plasma homocysteine (Hcy) 1 values of ϳ10 M for men and ϳ8 M for women are in the normal range. However, even a small increase (ϳ5 M) in total plasma Hcy is associated with a 60% increased risk of coronary artery disease for men and 80% for women (3). Moreover, a number of studies have demonstrated that smoking, excessive drinking of alcohol, obesity, type II diabetes, and an unhealthy diet contribute to mild hyperhomocysteinemia (1, 2). In addition, elevated plasma Hcy has recently been linked to Alzheimer's disease and cognitive impairment in the elderly (4, 5).Hcy is a non-protein amino acid derived from the catabolism of S-adenosylhomocysteine (AdoHcy), an immediate product of trans-methylation reactions that utilize S-adenosylmethionine (AdoMet) (6). Hcy has three possible fates: 1) methylation to methionine with N-5-methyltetrahydrofolate or betaine as the methyl donor, 2) conversion to cysteine via the trans-sulfuration pathway, and 3) release into extracellular fluids (e.g. plasma and urine). AdoMet-dependent methyltransferases catalyze many critical reactions including methylation of RNA, DNA, proteins, and small molecules such as guanidinoacetate and glycine (6). The potential of methyltransferases to regulate plasma Hcy is not well defined.Phosphatidylethanolamine N-methyltransferase (PEMT) is a liver-specific enzyme that generates AdoHcy during the conversion of one membrane lipid, phosphatidylethanolamine, into another membrane lipid, phosphatidylcholine (PC) (7). PEMT accounts for the formation of ϳ30% PC made in liver (8, 9). With this large capacity for PC synthesis and the generation of three AdoHcy molecules for each PC molecule synthesized, we hypothesized that the PEMT reaction might contribute significantly to Hcy in plasma. We have utilized the Pemt Ϫ/Ϫ mouse, hepatocytes derived from these mice, and overexpression of PEMT in McArdle RH7777 (rat hepatoma) cells to test this hypothesis. The results show that PEMT expression enhances plasma Hcy levels and the secretion of Hcy from hepatocyte...
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