CHANNELING OF INTERMEDIATES IN THE BIOSYNTHESIS OF PHOSPHATIDYLCHOLINE AND PHOSPHATIDYLETHANOLAMINE IN MAMMALIAN-CELLS

Previous studies with electropermeabilized cells have suggested the oc currence of metabolic compartmentation and Ca2+-dependent channeling o f intermediates of phosphatidylcholine (PC) biosynthesis in C6 rat gli oma cells. With a more accessible permeabilization technique, we inves tigated whether this is a more general phenomenon also occurring in ot her cell types and whether channeling is involved in phosphatidylethan olamine (PE) synthesis as well. C6 rat glioma cells, C3H10T1/2 fibrobl asts and rat hepatocytes were permeabilized with Staphylococcus aureus alpha-toxin. and the incorporation of the radiolabelled precursors ch oline, phosphocholine (P-choline), ethanolamine and phosphoethanolamin e (P-EA) into PC and PE were measured both at high and low Ca2+ concen trations. In glioma cells, permeabilization at high Ca2+ concentration did not affect [C-14]choline or [C-14]P-choline incorporation into PC . However, reduction of free Ca2+ in the medium from 1.8 mM to < 1 nM resulted in a dramatic increase in [C-14]P-choline incorporation into permeabilized cells, whereas [C-14]choline incorporation remained unaf fected. Also, in fibroblasts, reduction of extracellular Ca2+ increase d [C-14]P-choline and [C-14]P-EA incorporation into PC and PE respecti vely. In hepatocytes, a combination of cc-toxin and low Ca2+ concentra tion severely impaired [C-14]choline incorporation into PC. Therefore, alpha-toxin-permeabilized hepatocytes are not a good model in which t o study channeling of intermediates in PC biosynthesis. In conclusion, our results indicate that channeling is involved in PC synthesis in g lioma cells and fibroblasts. PE synthesis in fibroblasts is also at le ast partly dependent on channeling.