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13.
Sterol-mediated regulation of mevalonic acid synthesis
Plemenitaš, Ana, 1952- ; Havel, CM ; Watson, JA
Chinese hamster ovary-215 (CHO-215) mutant cells are auxotrophic for cholesterol. Berry and Chang (Berry, D. J., and Chang, T. Y. (1982) Biochemistry 21, 573-580) suggested that the metabolic lesion ...was at the levelof 4-methyl sterol oxidation. However, the observed cellular accumulationof lanosterol was not consistent with a defect at this metabolic site. With the use of a novel Silica Sep Pah sterol separation procedure, we demonstrated that 60-80 per cent of the acetonesoluble lipid radioactivity in (5-3H)mevalonate-labeled CHO-215 cells was incorporated into acidic sterols. 7(8),Cholesten-4a-methyl,4alpha-carboxy,3a-ol was the dominant end product. Inaddition to this acidic sterol, 7(8),24-cholestadien,4a-methyl,4alpha-carboxy,3a-ol and 7(8),24-cholestadien,4alpha-carboxy,3a-ol were also isolated. Incubation of cell-free extracts with (3H)7(8)cholesten-4a-methyl,4alpha-carboxy,3a-ol and pyridine nucleotides confirmed that CHO-215 4-carboxysterol decarboxylase activity was less than 1 per cent of that for wild type cells. Thus, a correspondence between decreased 4-carboxysterol decarboxylase activity and the spectrum of accumulated sterol products by intact CHO-215 cells was demonstrated. No detectable cholesterol was synthesized by CHO-215 cells. 3H-Product accumulation studies demonstrated that 7(8),24-cholestadien,4a-methyl,4alpha-carboxy,3a-ol increased prior to its subsequent saturation at the delta24 carbon. Furthermore, the steady state ratio for delta24-saturated acidic sterols/unsaturated acidic sterols was dependent on media cholesterol source and amount. Finally, the accumulated acidic sterol(s) were not regulatory signal molecules for the modulation of 3-hydroxy-8-methylglutaryl coenzyme. A reductase activity in response to cholesterol availability.
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