Mutant clone of Chinese hamster ovary cells lacking 3-hydroxy-3 -methylglutaryl coenzyme A reductase.
3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase) converts HMG-CoA to mevalonate, a key intermediate in the formation of cholesterol and several nonsterol isoprenoid compounds. Using the bromodeoxyuridine/bisbenzimide photosensitization technique, we isolated a mutant clone of Chinese hamster ovary cells that requires mevalonate for growth. This mutant, designated UT-2 cells, expresses 2-5% of the HMG-CoA reductase activity of parental Chinese hamster ovary cells, even after growth for 9 months in the absence of selective pressure. By immunoblotting, no cross-reactive HMG-CoA reductase protein was detected in UT-2 cells. Incorporation of [14C]acetate and [14C]pyruvate into cholesterol was less than 5% of that in parental Chinese hamster ovary cells. In contrast, [3H]mevalonate incorporation into cholesterol was normal. The activities of acetoacetyl-CoA thiolase and HMG-CoA synthase, the two enzymes that precede HMG-CoA reductase in the cholesterol biosynthetic pathway, were normal or slightly elevated in UT-2 cells. No gross deletions or rearrangements in the gene for HMG-CoA reductase were apparent when DNA from UT-2 cells was digested with restriction endonucleases, subjected to Southern blotting, and probed with a 32P-labeled cDNA for HMG-CoA reductase. We conclude that UT-2 cells have a mutation that specifically prevents the production of normal amounts of HMG-CoA reductase.[1]References
- Mutant clone of Chinese hamster ovary cells lacking 3-hydroxy-3 -methylglutaryl coenzyme A reductase. Mosley, S.T., Brown, M.S., Anderson, R.G., Goldstein, J.L. J. Biol. Chem. (1983) [Pubmed]
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