Localization of the membrane-associated CTP:phosphocholine cytidylyltransferase in Chinese hamster ovary cells with an altered membrane composition.
The subcellular localization of the membrane-associated CTP:phosphocholine cytidylyltransferase was determined in Chinese hamster ovary cells in which the phospholipid composition had been altered by growth in the presence of N-methylethanolamine or treatment with phospholipase C. Cell homogenates were fractionated on Percoll density gradients, and marker enzyme activities were used to determine the location of the cellular membrane fractions. The peak of cytidylyltransferase activity occurred in the gradient at a density intermediate to that of the peaks of endoplasmic reticulum and plasma membrane markers. The profile of cytidylyltransferase activity most closely resembled that of the Golgi membrane marker; however, upon sucrose gradient centrifugation, the profile of the Golgi apparatus was very different from that of cytidylyltransferase. Differential centrifugation suggested a nuclear membrane association of the enzyme. Cytidylyltransferase was associated with a membrane fraction that sedimented when subjected to very low speed centrifugation (65 x g, 5 min). From Percoll gradient fractions, nuclei were identified by microscopy, and they migrated with cytidylyltransferase activity. The data are consistent with a localization of cytidylyltransferase in the nuclear membrane.
Morand
J N
JN
Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907.
Kent
C
C
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002460:Cell Line; D002499:Centrifugation, Density Gradient / Q000379:methods; D019916:Choline-Phosphate Cytidylyltransferase; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D007700:Kinetics; D009685:Nuclear Envelope / Q000201:enzymology*; D009713:Nucleotidyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D010053:Ovary; D011205:Povidone; D012822:Silicon Dioxide; D013347:Subcellular Fractions / Q000201:enzymology / Q000648:ultrastructure; D013395:Sucrose
D013395:Sucrose; C016039:Percoll; D012822:Silicon Dioxide; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; D011205:Povidone