2 NEW SPHINGOMYELIN ANALOGS INHIBIT PHOSPHATIDYLCHOLINE BIOSYNTHESIS BY DECREASING MEMBRANE-BOUND CTP - PHOSPHOCHOLINE CYTIDYLYLTRANSFERASELEVELS IN HACAT CELLS

The effects of two newly synthesized sphingomyelin analogues on phosgh atidylcholine biosynthesis were investigated in the immortalized human keratinocyte cell line HaCaT. N-Acetyl-erythro -sphingosine-1-phospho choline (AcSM) and N-octanoyl-erythro-sphingosine-1-phosphocline (OcSM ) inhibited the incorporation of choline into phosphatidylcholine with half-inhibitory concentrations (IC50) of 6 mu/ml and 10 mu g/ml respe ctively. Further experiments revealed that AcSM and OcSM interfered wi th the translocation of the rate-limiting enzyme of phosphatidylcholin e biosynthesis, CTP:phosphocholine cytidylyltransferase (EC 2.7.7.15), in HaCaT cells and inhibited cytidylyltransferase activity in vitro. Despite the fact that OcSM was a potent inhibitor of cytidylyltransfer ase in vitro, its effects on phosphatidylcholine biosynthesis and tran slocation of cytidylyltransferase in HaCaT cells were less pronounced as compared with AcSM. Finally, we showed that the comparatively stron g effects of AcSM in cell culture experiments were due to the uptake o f large amounts of this sphingomyelin analogue into the cells. The res ults presented demonstrate that the activity of cytidylyltransferase m ay be negatively regulated by a high ratio of choline head group-conta ining sphingolipids.