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 production of recombinant proteins, namely, via the baculovirus vector expression system, little is known about the lipids in the cell membrane 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 amounts of ethanolamine plasmalogen. These phospholipids contain high proportions of monoenoic fatty acids, e.g., 16:1 and 18:1, which comprise more than 70% of the total fatty acids although small amounts of polyunsaturated fatty acids such as 18:2 and 20:4 are also present. When Sf9 cells were incubated in a culture medium containing [14C]oleic acid and [14C]arachidonic acid, a large portion of the labels were incorporated into membrane phospholipids. Using [14C]arachidonoyl-phospholipids as substrates for incubation with cell homogenate and subcellular fractions, results indicate the presence of a ca(2+)-independent phospholipase A (PLA2) in the Sf9 cell cytosol fraction. This PLA2 shows a high preference for hydrolysis 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 diacylglycerol (DAG) was released from [14C]arachidonoyl-PE in the Sf9 cells. Taken together, this study demonstrates active metabolism of membrane phospholipids in Sf9 cells, most likely mediated by acyltransferases and PLA2. Furthermore, despite the absence of PLC for PI, limited amount of DAG could be generated through hydrolysis of PE.
Yeh
L H
LH
Department of Chemical Engineering, University of Missouri, Columbia 65212, USA.
Bajpai
R K
RK
Sun
G Y
GY
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D003600:Cytosol / Q000201:enzymology; D004075:Diglycerides / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D005260:Female; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism*; D008861:Microsomes / Q000737:chemistry / Q000201:enzymology; D008928:Mitochondria / Q000737:chemistry / Q000201:enzymology; D019301:Oleic Acid / Q000378:metabolism; D010053:Ovary / Q000201:enzymology / Q000378:metabolism; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism*; D018411:Spodoptera / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D014280:Triglycerides / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
D004075:Diglycerides; D005227:Fatty Acids; D008563:Membrane Lipids; D010743:Phospholipids; D014280:Triglycerides; D016718:Arachidonic Acid; D019301:Oleic Acid; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D010738:Type C Phospholipases; D002118:Calcium