Phosphatidylcholine synthesis and

Diethanolamine has been shown to inhibit choline uptake into cultured Syrian hamster embryo (SHE) and Chinese hamster ovary cells and to inhibit the synthesis of phosphatidylcholine in in-vitro systems in a concentration-dependent, competitive and reversible manner (Lehman-McKeeman Gamsky, 1999, 2000). Diethanolamine treatment caused a marked reduction in hepatic choline metabolite concentrations in mice following two weeks of dermal dosing. The most pronounced reduction was in the hepatic concentration of phosphocholine, the intracellular storage form of choline (Stott et al, 2000). Moreover, the pattern by which choline metabolites were altered was similar to the pattern of change that has been observed following dietary choline deprivation in rodents (Pomfret et al, 1990). Excess choline also prevented diethanolamine-induced inhibition of phosphatidylcholine synthesis and incorporation of diethanolamine into SHE cell phospholipids (Lehman-McKeeman Gamsky, 2000). 

There are few commercial fungicides that have glycerophospholipid biosynthesis inhibition as their mode of action. The validated targets are phosphatidylcholine synthesis and phosphatidylinositol synthesis. 

Miquel, K., Pradines, A., Terce, F., Selmi, S., and Favre, G., 1998, Competitive inhibition of choline phosphotransferase by geranylgeraniol and famesol inhibits phosphatidylcholine synthesis and induces apoptosis in human lung adenocarcinoma A549 cells. JBiol Chem. 273 26179-26186... 

Vision E, Colombo C, Monti S, Giulidori P and Galli C (1998) S-adenosyl-L-methionine role in phosphatidylcholine synthesis and in vitro effects on the ethanol-induced alterations of lipid metabolism. Pharmacol Res 37 203-206. 

Acetylcholine Precursors. Early efforts to treat dementia using cholinomimetics focused on choline [62-49-7] (12) supplement therapy (Fig. 3). This therapy, analogous to L-dopa [59-92-7] therapy for Parkinson s disease, is based on the hypothesis that increasing the levels of choline in the brain bolsters acetylcholine (ACh) synthesis and thereby reverses deficits in cholinergic function. In addition, because choline is a precursor of phosphatidylcholine as well as ACh, its supplementation may be neuroprotective in conditions of choline deficit (104). 

The effect of Li+ upon the synthesis and release of acetylcholine in the brain is equivocal Li+ is reported to both inhibit and stimulate the synthesis of acetylcholine (reviewed by Wood et al. [162]). Li+ appears to have no effect on acetyl cholinesterase, the enzyme which catalyzes the hydrolysis of acetylcholine [163]. It has also been observed that the number of acetylcholine receptors in skeletal muscle is decreased by Li+ [164]. In the erythrocytes of patients on Li+, the concentration of choline is at least 10-fold higher than normal and the transport of choline is reduced [165] the effect of Li+ on choline transport in other cells is not known. A Li+-induced inhibition of either choline transport and/or the synthesis of acetylcholine could be responsible for the observed accumulation of choline in erythrocytes. This choline is probably derived from membrane phosphatidylcholine which is reportedly decreased in patients on Li+ [166],... 

Figure 1. Synthetic pathway for PS and PE in mammalian cells. The major steps occuring in the synthesis and interconversion of PS and PE are shown. The PS synthases condense serine with a phosphatidyl moiety derived from PC and PE. The nascent PS can be converted to PE by decarboxylation. PE can also be formed by transfer of a phosphoethanolamine moiety from CDP-ethanolamine to diacylglycerol via the Kennedy pathway. The abbreviations used are PC, phosphatidylcholine PS, phosphatidylserine PE, phosphatidylethanolamine DG, diacylglycerol PSD, phosphatidylserine decarboxylase PSS, PS synthase.

Figure 1. Pathways for the synthesis of phosphatidylcholine, phosphatidylethanolamine and sphingomyelin. Abbreviations CK, choline kinase CPT, cholinephosphotransferase CT, CTP phosphooholine cytidylyltransferase DAG, diacylglycerol PC, phosphatidylcholine PE, phosphatidylethanolamine PEMT, phosphatidylethanolamine-N-methyltransferase SM, sphingomyelin SMase, sphingomyelinase SMsyn, sphingomyelin synthase.

Figure 2. Effect of Ca-ceramide on phosphatidylcholine and phosphatidylethanolamine synthesis in rat-2 fibroblasts. Cells were treated for 2 h in the absence (open bars) or presence (hatched bars) of 25pM C6-ceramide and the following parameters were determined i) the incorporation of [ H]choline and [ H]ethanolamine into phosphatidylcholine (PC) and phosphatidylethanolamine (PE), respectively (panel A) and in CDP-choline and CDP-ethanolamine, respectively (panel B) and ii) the in vitro activity of choline- and ethanolaminephosphotransferase (CPT and EPT) (panel C).

Wright, M.M., Henneberry, A.L., Lagace, T.A., Ridgway, N.D., and McMaster, C.R., 2001, Uncouphng Famesol Induced Apoptosis from its Inhibition of Phosphatidylcholine Synthesis. . Biol Chem. 276 25254-25261. 

Glycerophospholipids are used for membrane synthesis and for producing a hydrophilic surface layer on lipoproteins such as VLDL. In cell membranes, they also serve as a reservoir of second messengers such as diacylglycerol, inositol 1,4,5-triphosphate, and arachidonic acid. Their structure is similar to triglycerides, except that the last fatty acid is replaced by phosphate and a water-soluble group such as choline (phosphatidylcholine, lecithin) or inositol (phosphatidyl-inositol). 

Lehman-McKeeman, L.D. Gamsky, E.A. (1999) Diethanolamine inhibits choline uptake and phosphatidylcholine synthesis in Chinese hamster ovary cells. Biochem. biophys. Res. Comm., 262, 600-604... 

The phospholipids in milk are synthesized by the mammary cell via pathways that are common to other mammalian cells. For further information on the synthesis of phospholipids in the mammary cell, see Kinsella and Infante (1978) and Patton and Jensen (1976). The major glycerophospholipids are phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, and phosphatidylinositol. A more complete composition is given in Table 4.6, Patton and Jensen (1976). The acyl and alkyl compositions will be given later. In milk, the glycerophospholipids are found predominantly in the diacyl form. However, small... 

Phospholipids have been shown to exchange between different membranes, e.g., of mitochondria and the ER. Exchanges of phosphatidylcholine, phosphatidylinositol, and sphingomyelin are catalyzed by specific exchange proteins (Box 21-A).238 2383 These proteins may also participate in signaling, but their major function may be to transport the phospholipids from their sites of synthesis to the various membranes of the cell. 

Lipids also show asymmetrical distributions between the inner and outer leaflets of the bilayer. In the erythrocyte plasma membrane, most of the phosphatidylethanolamine and phosphatidylserine are in the inner leaflet, whereas the phosphatidylcholine and sphingomyelin are located mainly in the outer leaflet. A similar asymmetry is seen even in artificial liposomes prepared from mixtures of phospholipids. In liposomes containing a mixture of phosphatidylethanolamine and phosphatidylcholine, phosphatidylethanolamine localizes preferentially in the inner leaflet, and phosphatidylcholine in the outer. For the most part, the asymmetrical distributions of lipids probably reflect packing forces determined by the different curvatures of the inner and outer surfaces of the bilayer. By contrast, the disposition of membrane proteins reflects the mechanism of protein synthesis and insertion into the membrane. We return to this topic in chapter 29. 

The energy state of the cell dictates the relative rates of phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol biosynthesis. When energy is in short supply, the level of cAMP rises leading to inhibition of fatty acid biosynthesis (see chapter 18). This in turn decreases the supply of diacylglycerol, which limits the synthesis of phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol. When sufficient diacylglycerol is pres-... 

In some cases the functions of phospholipases in cells are purely degradative and result in the release of the phospholipid components (fatty acids, glycerol, phosphate, and head-groups). But in many cases phospholipases have important roles in synthesis and regulation. For example, we have seen how phospholipase A2 catalyzes the first step in the remodeling of phosphatidylcholine to the surfactant... 

Boggs K. P., Rock C. O., and Jackowski S. (1995). Lysophosphatidylcholine and l-O-octadecyl-2-O- methyl-rac-glycero-3-phosphocholine inhibit the CDP-choline pathway of phosphatidylcholine synthesis at the CTP phosphocholine cytidylyltransferase step. J. Biol. Chem. 270 7757-7764. 

Gasull T., DeGregorio-Rocasolano N., Zapata A., and Trullas R. (2000). Choline release and inhibition of phosphatidylcholine synthesis precede excitotoxic neuronal death but not neurotoxicity induced by serum deprivation. J. Biol. Chem. 275 18350-18357. 

To investigate the specificity of DGAT inhibitors, synthesis of lipids including TG, phosphatidylcholine (PC), and phosphatidylethanolamine (PE) was measured in an intact cell assay using Raji cells by our established method [54], Raji cells are incubated in the presence of [14C]oleic acid with or without a test sample in a total volume of 0.2 ml. After a 20-min incubation at 37°C, [14C]oleic acid is incorporated mainly into PC, PE, and TG, which are extracted and separated by TLC. The distribution of radioactivity in these lipids on TLC is analyzed. 

Synthetic Schemes. There are basically two main routes to preparation of mixed acid (enantiomeric) diacylphosphatidylcholines. The first involves a total de novo synthesis and the second centers on a partial or semisynthesis using a highly purified naturally occurring phosphatidylcholine as starting material. Both approaches have merits and drawbacks, which are noted as follows. 

Partial (Semi) Synthesis. A blend of two methodologies can be used quite effectively to prepare mixed acid phosphatidylcholines, which are of prime importance in elucidating the structure of naturally occurring phospho-glyceride and phospholipase A2 activity. Perhaps the best scenario would be to illustrate the activity of the phospholipase A2 (from Crotalus adamanteus snake venom) toward a racemic phosphatidylcholine sample and toward individual sn-1 and sn-3 enantiomers. In each case the same result would be found, and so only the racemic mixture reaction is depicted in Figure 4-8. 

Griac, P., and Henry, S.A., 1996, Phosphatidylcholine biosynthesis in Saccharomyces cerevisiae Effects on regulation of phospholipid synthesis and respiratory competence. In Op den Kamp, J.A.F. (ed.), NATO ASI Series Molecular Dynamics of Biological Membranes. Springer, Verlag, pp. 339-346. 

Choline can be used for synthesis of phosphatidylcholine by reaction between CDP-choline and diacylglycerol. Under normal conditions, the major pathway of phosphatidylcholine synthesis is by the incorporation of... 

---