CLONING AND EXPRESSION OF MOUSE-LIVER PHOSPHATIDYLSERINE SYNTHASE-1 CDNA - OVEREXPRESSION IN RAT HEPATOMA-CELLS INHIBITS THE CDP-ETHANOLAMINE PATHWAY FOR PHOSPHATIDYLETHANOLAMINE BIOSYNTHESIS

In eukaryotic cells, phosphatidylserine (PtdSer) is synthesized by two distinct synthases on the endoplasmic reticulum by a base-exchange re action in which the polar head-group of an existing phospholipid is re placed with serine. We report the cloning and expression of a cDNA for mouse liver PtdSer synthase-1, The deduced protein sequence is >90% i dentical to that of PtdSer synthase-1 from Chinese hamster ovary cells and a sequence from a human myeloblast cell Line, PtdSer synthase-1 c DNA was stably expressed in M.9.1.1 cells which are mutant Chinese ham ster ovary cells defective in PtdSer synthase-1 activity, are ethanola mine auxotrophs, and have a reduced content of PtdSer and phosphatidyl ethanolamine (PtdEtn), The growth defect of M.9.1.1 cells was eliminat ed, and a normal phospholipid composition was restored in the absence of exogenous ethanolamine, implying that the cloned cDNA encoded PtdSe r synthase, Mouse liver PtdSer synthase-1 was also expressed in McArdl e 7777 rat hepatoma cells, In addition to a 3-fold higher in vitro ser ine-exchange activity, these cells also exhibited enhanced choline- an d ethanolamine-exchange activities and incorporated more [H-3]serine i nto PtdSer than did control cells. However, the levels of PtdSer and P tdEtn in cells overexpressing PtdSer synthase-1 activity were not incr eased, Excess PtdSer produced by the transfected cells was rapidly dec arboxylated to PtdEtn and the degradation of PtdSer, and/or PtdEtn der ived from PtdSer, was increased, Moreover, the CDP-ethanolamine pathwa y for PtdEtn biosynthesis was inhibited, These data suggest that (i) c ellular levels of PtdSer and PtdEtn are tightly controlled, and (ii) t he metabolism of PtdSer and PtdEtn is coordinately regulated to mainta in phospholipid homeostasis.