MEMBRANE LIPID-METABOLISM AND PHOSPHOLIPASE-ACTIVITY IN INSECT SPODOPTERA-FRUGIPERDA-9 OVARIAN-CELLS

Although there is increasing use of insect ovarian Sf9 cells for the p roduction of recombinant proteins, namely, via the baculovirus vector expression system, little is known about the lipids in the cell membra ne and whether endogenous phospholipases are present for regulation of the cell membrane lipids. In this study, analysis of membrane lipids of Sf9 cells indicated the presence of phosphatidylethanolamine (PE) ( diacyl type) and phosphatidylcholine as major phospholipids, followed by phosphatidylserine and phosphatidylinositol (PI), and only trace am ounts of ethanolamine plasmalogen. These phospholipids contain high pr oportions of monoenoic fatty acids, e.g., 16:1 and 18:1, which compris e more than 70% of the total fatty acids although small amounts of pol yunsaturated fatty acids such as 18:2 and 20:4 are also present. When Sf9 cells were incubated in a culture medium containing [C-14]oleic ac id and [C-14]arachidonic acid, a large portion of the labels were inco rporated into membrane phospholipids. Using [C-14]arachidonoyl-phospho lipids as substrates for incubation with cell homogenate and subcellul ar fractions, results indicate the presence of a Ca2+-independent phos pholipase A (PLA(2)) in the Sf9 cell cytosol fraction. This PLA(2) sho ws a high preference for hy drolysis of PE and is active at a pH range of 7-9. Unlike the brain cells which contain active phospholipase C ( PLC) specific for phosphatidylinositol, only limited amount of diacylg lycerol (DAG) was released from [C-14]arachidonoyl-PE in the Sf9 cells . Taken together, this study demonstrates active metabolism of membran e phospholipids in Sf9 cells, most likely mediated by acyltransferases and PLA(2). Furthermore, despite the absence of PLC for PI, limited a mount of DAG could be generated through hydrolysis of PE.