Appearance of crystalloid endoplasmic reticulum in compactin-resistant Chinese hamster cells with a 500-fold increase in 3-hydroxy-3-methylglutaryl-coenzyme A reductase.
We have developed a line of Chinese hamster ovary cells with a 500-fold increase in 3-hydroxy-3-methylglutaryl-coenzyme A reductase, the membrane-bound enzyme that controls cholesterol synthesis. This line, designated (UT-1, was obtained by stepwise adaptation of cells to growth in increasing concentrations of compactin, a competitive inhibitor of reductase. Reductase accounts for approximately 2% of total cell protein in UT-1 cells, as calculated from enzyme specific activity and by immunoprecipitation of reductase after growth of cells in [35S]methionine. After solubilization in the presence of the protease inhibitor leupeptin and electrophoresis in NaDodSO4/polyacrylamide gels, reductase subunits from UT-1 cells were visualized by immunoblotting as a single band (Mr = 62,000). To accommodate the increased amounts of reductase, UT-1 cells developed marked proliferation of tubular smooth endoplasmic reticulum (ER) membranes, as revealed by immunofluorescence and electron microscopy. The ER tubules were packed in crystalloid hexagonal arrays. When UT-1 cells were incubated with low density lipoprotein, reductase activity was suppressed by 90% in 12 hr and the crystalloid ER disappeared. UT-1 cells should be useful for studies of the regulation of reductase and also for studies of the synthesis and degradation of smooth ER.
D000818:Animals; D002460:Cell Line; D002784:Cholesterol / Q000378:metabolism; D002999:Clone Cells; D006224:Cricetinae; D003470:Culture Media; D004351:Drug Resistance; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure*; D005260:Female; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000494:pharmacology; D008148:Lovastatin / Q000031:analogs & derivatives*; D009281:Naphthalenes / Q000494:pharmacology*; D010053:Ovary
D003470:Culture Media; D008077:Lipoproteins, LDL; D009281:Naphthalenes; C012258:mevastatin; D002784:Cholesterol; D008148:Lovastatin; D006903:Hydroxymethylglutaryl CoA Reductases