Regulation of de novo phosphatidylinositol synthesis

Abstract

Mechanisms that function to regulate the rate of de novo phosphatidylinositol (PtdIns) synthesis in mammalian cells have not been elucidated. In this study, we characterize the effect of phorbol ester treatment on de novo PtdIns synthesis in C3A human hepatoma cells. Incubation of cells with 12-O-tetradecanoyl phorbol 13-acetate (TPA) initially (1-6 h) results in a decrease in precursor incorporation into PtdIns; however, at later times (18-24 h), a marked increase is observed. TPA-induced glucose uptake from the medium is not required for observation of the stimulation of PtdIns synthesis, because the effect is apparent in glucose-free medium. Inhibition of the activation of arachidonic acid substantially blocks the synthesis of PtdIns but has no effect on the synthesis of phosphatidylcholine (PtdCho). Increasing the concentration of cellular phosphatidic acid by blocking its conversion to diacylglycerol, on the other hand, enhances the synthesis of PtdIns and inhibits the synthesis of PtdCho. The TPA-induced stimulation of PtdIns synthesis is not the result of the concomitant TPA-induced G1 arrest, because G1 arrest induced by mevastatin has no effect on PtdIns synthesis. Inhibition of protein kinase C activity blocks the stimulatory action of TPA on de novo synthesis of PtdIns but has no effect on TPA-induced inhibition.Potential sites of enzymatic regulation are discussed.-Nuwayhid, S. J., M. Vega, P. D. Walden, and M. E. Monaco. Phosphatidylinositol (PtdIns) is an essential acidic phospholipid found in the membranes of mammalian cells (1) as well as on the chromatin (2). A single molecular species composed of stearate (R9) and arachidonate (R99) accounts for z80% of the total PtdIns. There are two pathways for the synthesis of PtdIns: the de novo pathway and the salvage pathway, which converge at the level of phosphatidic acid (PtdOH). The de novo pathway uses dihydroxyacetone phosphate derived from glucose to make glycerol phosphate, which is subsequently acylated at the sn-1 position with stearate and then at the sn-2 position with arachidonate to yield PtdOH. In the salvage pathway, diacylglycerol (DAG), derived from phospholipase C-mediated hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdInsP 2 ), is converted to PtdOH through the action of DAG kinase. In the first instance, there is a net increase in PtdIns, whereas in the second, there is not. The central element of both pathways is the production of PtdOH. For the salvage pathway, it appears that the conversion of DAG to PtdOH by DAG kinase is stimulated as a result of agonist-induced polyphosphoinositide turnover. Whether this results from an increase in DAG substrate and/or an increase in DAG kinase enzyme activity is unclear. This laboratory has previously demonstrated that the two pathways can be differentiated in situ on the basis of the inhibition by triacsin C, a specific inhibitor of arachidonate-specific long-chain fatty acyl-CoA synthetase (ACSL4) in situ (3, 4).Although regulation of the salvage pathway via the activation ...

Transport and Metabolism of Radiolabeled Choline in Hepatocellular Carcinoma

“…The rate of PC biosynthesis does not appear to be influenced significantly by the rate of Cho transport in most nonproliferating cells such as rat liver and rat kidney cortex 54–55. In addition, for these nonproliferating cells, Cho is metabolized through both the CDP-Cho pathway and the betaine oxidation pathway.…”

Fatty acid metabolism in breast cancer subtypes

“…Triacsin C is a fatty acid analogue that has been shown to inhibit ACSL 1, ACSL3 and ACSL4. This compound causes apoptosis of many cancer cell types, including breast [71, 110–113], and has been demonstrated to inhibit the growth of xenograft tumors [110]. Lipid droplets comprise an emerging target in a number of diseases, including cancer, and it has been suggested that the lipid droplet protein, PLIN2, might make an effective target [114].…”

Lipids in Entamoeba histolytica: Host-Dependence and Virulence Factors

“…This synthase produces L-myo-inositol 1-phosphate by the transformation of D-glucose-6-phosphate (Lohia et al, 1999). During de novo PI synthesis in eukaryotes, myoinositol-phosphate is dephosphorylated by inositol 3-phosphate monophosphatase and transformed into myo-inositol, which is converted in PI with CDP-diacylglycerol in a reaction catalyzed by PI synthase (also called CDP-diacylglycerol-inositol 3-phosphatidyltransferase; Nuwayhid et al, 2006). In the E. histolytica genome, there is one CDP-diacylglycerol-inositol 3phosphatidyltransferase (EHI_069630), an enzyme related to the synthesis of CDP-diacylglycerol, suggesting that PI synthesis pathway is conserved in this parasite.…”