Regulation of phosphatidylcholine biosynthesis in Chinese hamster ovary cells by reversible membrane association of CTP: phosphocholine cytidylyltransferase.
Treatment of Chinese hamster ovary cells with phospholipase C was previously shown to stimulate the CDP-choline pathway for phosphatidylcholine biosynthesis, and to cause activation of the CTP:phosphocholine cytidylyltransferase with a concomitant change in subcellular location of the enzyme (Sleight, R., and Kent, C. (1983) J. Biol. Chem. 258, 831-835). This paper presents a detailed analysis of the early events in the phospholipase C treatment, and provides evidence that the increased cytidylyltransferase activity causes the increased flux through the pathway. The time courses for the increase in cytidylyltransferase activity, increase in amount of membrane-associated enzyme, decrease in phosphocholine levels, and increase in phosphatidylcholine synthesis were similar, with all changes occurring within 30 min after addition of phospholipase C. These events preceded a decrease in cellular choline levels which correlated with a decreased capacity for choline uptake. The rate at which radioactive label was lost from pulse-labeled phosphocholine was the same as the rate at which label was incorporated into phosphatidylcholine, and these rates were stimulated 2.2-fold by phospholipase C treatment. We have also shown that the association of cytidylyltransferase with membranes was rapidly reversible when phospholipase C was removed from the cultures, and that the rate of decrease in phosphatidylcholine synthesis paralleled the rate of decrease in cytidylyltransferase activity. Cytidylyltransferase became reassociated with membranes when phospholipase C was added back to cultures from which it was previously removed. These results represent the first detailed account of the time frame involved in regulating phosphatidylcholine synthesis by the reversible association of cytidylyltransferase with cellular membranes.
Wright
P S
PS
Morand
J N
JN
Kent
C
C
D000818:Animals; D002460:Cell Line; D002794:Choline / Q000378:metabolism; D019916:Choline-Phosphate Cytidylyltransferase; D006224:Cricetinae; D003412:Cricetulus; D003513:Cycloheximide / Q000494:pharmacology; D005260:Female; D009713:Nucleotidyltransferases / Q000378:metabolism*; D010053:Ovary / Q000187:drug effects / Q000201:enzymology*; D010713:Phosphatidylcholines / Q000096:biosynthesis*; D010767:Phosphorylcholine / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
D010713:Phosphatidylcholines; D010767:Phosphorylcholine; D003513:Cycloheximide; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; D010738:Type C Phospholipases; D002794:Choline