Cell-free transfer of phospholipids between the endoplasmic reticulum and the Golgi apparatus of leek seedlings

Abstract

The transfer of lipids between the endoplasmic reticulum and the Golgi apparatus was investigated in vitro using a cell-free system from leek seedlings. Lipids of the donor membranes (endoplasmic reticulum) were radiolabeled either by incubating leek seedlings with [1- 14C]acetate or [ 3H]acetate. Acceptor membranes (Golgi apparatus) were unlabeled and immobilized on nitrocellulose strips. The assay measured the lipid transfer resulting from both an ATP-independent process and an ATP- and cytosol-dependent process. A significant ATP- and cytosol-dependent lipid transfer was observed only in the case of the endoplasmic reticulum as donor and the Golgi apparatus as acceptor. Lipids transferred in an ATP-dependent manner were chiefly phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine. The stimulation of lipid transfer by ATP as compared to the ATP-independent process was + 79% (PC), + 123% (PS) and + 69% (PE). On the other hand, PI was not transferred in an ATP-dependent manner (the stimulation by ATP was only 20%). This supports the theory that a sorting of phospholipids takes place in the donor membrane. Moreover, a formation of lysoPC was observed only in the presence of ATP (+ 330%). The ATP-dependent lipid transfer was inhibited by N- ethylmaleimide , indicating the involvement of cytosolic (but no phospholipid transfer proteins) or membrane proteins in the transfer process. The ATP-dependent transfer of lipids was also diminished at 12°C showing the sensitivity to low temperatures of the transfer of lipids between the endoplasmic reticulum and the Golgi apparatus.