A 25-hydroxycholesterol-resistant cell line deficient in acyl-CoA: cholesterol acyltransferase.
We describe a line of mutant Chinese hamster ovary cells, SRD-4 cells, that lacks acyl-coenzyme A:cholesterol acyltransferase (ACAT) activity and fails to synthesize cholesteryl esters when stimulated with 25-hydroxycholesterol or low density lipoprotein. The cells also have a partial defect in their ability to repress transcription of three sterol-regulated genes, 3-hydroxy-3-methylglutaryl-coenzyme A synthase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, and the low density lipoprotein receptor. The cells were selected by mutagenesis followed by growth in the presence of 25-hydroxycholesterol, which kills the parental cells by cholesterol depletion, owing to an inhibition of cholesterol synthesis and a stimulation of cholesterol esterification. Treatment of parental cells with compound 58-035 (3-(decyldimethylsilyl)-N-[2-(4-methylphenyl)-1- phenylethyl]propanamide), an inhibitor of ACAT, abolished cholesterol esterification but did not reproduce the defect in gene repression seen in the SRD-4 cells, and it only partially reproduced the resistance to the killing effect of 25-hydroxycholesterol. We conclude that the SRD-4 cells most likely have two independent defects, one in ACAT and the other in a factor that mediates sterol-dependent transcriptional repression. The SRD-4 cells thus resemble a line of hamster cells previously isolated (Cadigan, K.M., Heider, J.G., and Chang, T.-Y. (1988) J. Biol. Chem. 263, 274-282), which has similar independent defects. The results raise the possibility that a partial resistance to sterol repression provides a growth advantage to cells that lack ACAT.
Metherall
J E
JE
University of Texas Southwestern Medical Center, Department of Molecular Genetics, Dallas 75235.
Ridgway
N D
ND
Dawson
P A
PA
Goldstein
J L
JL
Brown
M S
MS
D000577:Amides / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002788:Cholesterol Esters / Q000096:biosynthesis; D006224:Cricetinae; D003412:Cricetulus; D006706:Homeostasis; D006888:Hydroxycholesterols / Q000378:metabolism*; D006904:Hydroxymethylglutaryl-CoA Synthase / Q000378:metabolism; D009154:Mutation; D017646:Organosilicon Compounds; D012313:RNA / Q000032:analysis; D011973:Receptors, LDL / Q000378:metabolism; D002785:Sterol O-Acyltransferase / Q000172:deficiency*; D013261:Sterols / Q000378:metabolism
D000577:Amides; D002788:Cholesterol Esters; D006888:Hydroxycholesterols; D017646:Organosilicon Compounds; D011973:Receptors, LDL; D013261:Sterols; D012313:RNA; C040256:SAN 58035; D002785:Sterol O-Acyltransferase; D006904:Hydroxymethylglutaryl-CoA Synthase