Uptake of Exogenous PhosphatidyJserine by Human Neuroblastoma Cells Stimulates the Incorporation of [methyl‐l4C]Cholne into Phosphatidylcholine

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Abstract
The phosphatidylserine (PtdSer) content of human cholinergic neuroblastoma (LA‐N‐2) cells was manipulated by exposing the cells to exogenous PtdSer, and the effects on phospholipid content, membrane composition, and incorporation of choline into phosphatidylcholine (PtdCho) were investigated. The presence of liposomes containing PtdSer (10‐130 μM) in the medium caused time‐ and concentration‐dependent increases in the PtdSer content of the cells, and smaller and slower increases in the contents of other membrane phospholipids. The PtdSer levels in plasma membrane and mitochondrial fractions prepared by discontinuous sucrose density gradient centrifugation increased by 50 and 100%, respectively, above those in control cells after 24 h of exposure to PtdSer (130 μM). PtdSer caused a concomitant, concentration‐dependent increase of up to twofold in the incorporation of [methyl‐14C]choline chloride into PtdCho at a choline concentration (8.5 μM) compatible with activation of the CDP‐choline pathway, suggesting that the levels of PtdSer in membranes may serve as a stimulus to regulate overall membrane composition. PtdSer caused a mean increase of 41% in PtdCho labeling, but the phorbol ester, phorbol 12‐myristate 13‐acetate (PMA), which stimulates PtdCho synthesis in a number of cell lines, increased [4C]PtdCho levels by only 14% in LA‐N‐2 cells, at a concentration (100 nM) which caused complete translocation of the calcium‐ and phospholipid‐dependent enzyme protein kinase C to the membrane. The translocation was inhibited by prior exposure of the cells to PtdSer. Treatment with PMA for 24 h diminished protein kinase C activity by 80%, but increased the labeling of PtdCho in both untreated and PtdSer‐treated cells. These data suggest that uptake of PtdSer by LA‐N‐2 cells alters both the phospholipid composition of the membrane and synthesis of the major membrane phospholipid PtdCho; the latter effect does not involve activation of protein kinase C.

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