Lack of correlation between 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and lovastatin resistance in nerve growth factor treated PC-12 cells
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1. The relationships among the mevalonic acid (MVA) forming enzyme, 3-hydroxy-3-methylglutaryl coenzyme A (CoA) reductase, cell growth and differentiation, and the cytotoxic effects of the reductase inhibitor lovastatin were studied in PC-12 cells, exposed to growth factors.
2. When added individually, nerve growth factor (NGF), basic fibroblast growth factor, and epidermal growth factor induce an increase in HMG-CoA reductase activity in cells grown in serum-containing medium. In the presence of serum, the effect of NGF on HMG-CoA reductase is persistent.
3. Short-term serum starvation and long-term NGF treatment, in combination, have an additive effect, resulting in a high reductase activity.
4. Unlike serum and MVA, which downregulate levels of HMG-CoA reductase by accelerating its degradation, NGF upregulates reductase by slowing the rate of its degradation. This mechanism, however, appears to operate only in the presence of serum, as after prolonged growth with NGF in serum-free medium, cells have a low reductase activity.
5. PC-12 cells grown in the absence of NGF are highly sensitive to lovastatin (25 µM) and more than 70% of the cells die after 48 hr. NGF confers lovastatin resistance on cells grown in the presence or in the absence of serum (only 30–40% cell death after 48 hr with lovastatin).
6. NGF-induced resistance on lovastatin develops with time and is apparent only in the well-differentiated PC-12 cells whether or not the cells express a high reductase activity.
7. Thus, levels of HMG-CoA reductase activity and lovastatin resistance in PC-12 cells are not directly correlated, though clearly inversed lovastatin cytotoxicity and elevated reductase activities are expressed during the period of cell proliferation.
8. These data suggest that fully differentiated neuronal cells may not be affected by prolonged high doses of lovastatin.
Key wordsPC12 nerve growth factor 3-hydroxy-3-methylglutaryl coenzyme reductase lovastatin
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- Alberts, A. W., Chen, J., Kuron, G., Hunt, V., Huff, J., Hoffman, C., Rothrock, J., Lopez, M., Joshua, H., Harris, E., Patchett, A., Monaghan, R., Currie, S., Stapley, E., Albers-Schonberg, G., Hensens, O., Hirshfield, J., Hoogsteen, K., Liesch, J., and Springer, J. (1980). Mevinolin: A highly potent competitive inhibitor of hydroxymethylglutaryl-coenzyme A reductase and a cholesterol-lowering agent.Proc. Natl. Acad. Sci. USA 773957–3961.Google Scholar
- Batistatou, A., and Greene, L. A. (1991). Aurintricarboxylic acid redcues PC12 cells and sympathetic neurons from cell death caused by nerve growth factor deprivation: Correlation with suppression of endonuclease activity.J. Cell Biol. 115461–471.Google Scholar
- Brown, M. S., Faust, J. R., and Goldstein, J. L. (1978). Induction of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in human fibroblasts incubated with compactin (ML-236B), a competitive inhibitor of the reductase.J. Biol. Chem. 2531121–1128.Google Scholar
- Casey, P. J. (1992). Biochemistry of protein prenylation.J. Lipid Res. 331731–1740.Google Scholar
- Cuthbert, J. A., Russell, D. W., and Lipsky, P. E. (1989). Regulation of low density lipoprotein receptor gene expression in human lymphocytes.J. Biol. Chem. 2641298–1304.Google Scholar
- Endo, A., and Hasumi, K. (1989). Biochemical aspect of HMG-CoA reductase inhibitors.Adv. Enz. Reg. 2753–64.Google Scholar
- Fenton, R. G., Kung, H., Longo, D. L., and Smith, M. R. (1992). Regulation of intracellular actin polymerization by prenylated cellular proteins.J. Cell Biol. 117347–356.Google Scholar
- Goldstein, J. L., and Brown, M. S. (1990). Regulation of the mevalonate pathway.Nature 343425–430.Google Scholar
- Greene, L. A. (1978). Nerve growth factor prevents the death and stimulates the neuronal differentiation of clonal PC12 pheochromocytoma cells in serum-free medium.J. Cell Biol. 78747–755.Google Scholar
- Haklai, R., and Kloog, Y. (1991). Relationship among methylation, isoprenylation, and GTP binding in 21- to 23-kDa proteins of neuroblastoma.Cell. Mol. Neurobiol. 11415–433.Google Scholar
- Haklai, R., Lerner, S., and Kloog, Y. (1993). Nerve growth factor induces a succession of increases in isoprenylated methylated small GTP-binding proteins of pheochromocytoma cells.Neuropeptides 2411–25.Google Scholar
- Halegoua, S., Armstrong, R. C., and Kremer, N. E. (1991). Dissecting the mode of action of a neuronal growth factor.Curr. Top. Microbiol. Immunol. 165119–170.Google Scholar
- Jakobisiak, M., Bruno, S., Skierski, J. S., and Darzynkiewicz, Z. (1991). Cell cycle-specific effects of lovastatin.Proc. Natl. Acad. Sci. USA 883628–3632.Google Scholar
- Keyomarsi, K., Sandoval, L., Band, V., and Pardee, A. B. (1991). Synchronization of tumor and normal cells from G1 to multiple cell cycles by lovastatin.Cancer Res. 513602–3609.Google Scholar
- Kita, T., Brown, M. S., and Goldstein, J. L. (1980). Feedback regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase in livers of mice treated with Mevinolin, a competitive inhibitor of the reductase.J. Clin. Invest. 661094–1100.Google Scholar
- Kremer, N. E., D'Arcangelo, G., Thomas, S. M., DeMarco, M., Brugge, J. S., and Halegoua, S. (1991). Signal transduction by nerve growth factor and fibroblast growth factor in PC-12 cells requires a sequence of src and ras actions.J. Cell Biol. 115809–819.Google Scholar
- Langan, T. J., and Slater, M. C. (1991a). Cell cycling of astrocytes and their precursors in primary cultures; A mevalonate requirement identified in late G1, but before the G1/S transition, involves polypeptides.J. Neurochem. 561058–1068.Google Scholar
- Langan, T. J., and Slater, M. C. (1991b). Quiescent astroglia in long-term primary cultures re-enter the cell cycle and require a non-sterol isoprenoid in late G1.Brain Res. 5489–17.Google Scholar
- Lerner, S., Haklai, R., and Kloog, Y. (1992). Isoprenylation and carboxymethylation in small GTP-binding proteins of pheochromocytoma (PC-12) cells.Cell. Mol. Neurobiol. 12333–351.Google Scholar
- Maltese, W. A., and Sheridan, K. M. (1987). Isoprenylated proteins in cultured cells: Subcellular distribution and changes related to altered morphology and growth arrest induced by mevalonate deprivation.J. Cell. Physiol. 133471–481.Google Scholar
- Mosmann, T. (1983). Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays.J. Immunol. Methods 6555–63.Google Scholar
- Muroya, K., Hattori, S., and Nakamura, S. (1992). Nerve growth factor induces rapid accumulation of the GTP-bound form of p21ras in rat pheochromocytoma PC12 cells.Oncogene 7277–281.Google Scholar
- Nakanishi, M., Goldstein, J. L., and Brown, M. S. (1988). Multivalent control of 3-hydroxy-3-methylglutaryl coenzyme A reductase.J. Biol. Chem. 2638929–8937.Google Scholar
- Qui, M. S., Pitts, A. F., Winters, T. R., and Green, S. H. (1991). Ras isoprenylation is required for ras-induced but not for NGF-induced neuronal differentiation of PC12 cells.J. Cell. Biol. 115795–808.Google Scholar
- Roitelman, J., and Simoni, R. D. (1992). Distinct sterol and nonsterol signals for the regulated degradation of 3-hydroxy-3-methylglutaryl-CoA reductase.J. Biol. Chem. 26725264–25273.Google Scholar
- Roth, M., Emmons, L. R., Perruchoud, A., and Block, L. H. (1991). Expression of low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl coenzyme A reductase genes are stimulated by recombinant platelet-derived growth factor isomers.Proc. Natl. Acad. Sci. USA 881888–1892.Google Scholar
- Rukenstein, A., Rydel, R. E., and Greene, L. A. (1991). Multiple agents rescue PC12 cells from serum-free cell death by translation- and transcription-independent mechanisms.J. Neurosci. 112552–2563.Google Scholar
- Rydel, R. E., and Greene, L. A. (1987). Acidic and basic fibroblast growth factors promote stable neurite outgrowth and neuronal differentiation in cultures of PC12 cells.J. Neurosci. 73639–3653.Google Scholar
- Schmidt, R. A., Glomset, J. A., Wight, T. N., Habenicht, A. J. R., and Ross, R. (1982). A study of the influence of mevalonic acid and its metabolites on the morphology of Swiss 3T3 cells.J. Cell Biol. 95144–153.Google Scholar
- Shapiro, D. J., Nordstrom, J. L., Mitschelan, J. J., Rowele, V. W., and Schmike, R. T. (1974). Micro assay for 3-hydroxy-3-methylglutaryl CoA reductase in rat liver and in L-cell fibroblasts.Biochim. Biophys. Acta 370369–377.Google Scholar
- Sinesky, M., Beck, L. A., Leonard, S., and Evans, R. (1990). Differential inhibitory effects of lovastatin on protein isoprenylation and sterol synthesis.J. Biol. Chem. 26519937–19941.Google Scholar
- Siperstein, M. D. (1984). Role of cholesterogenesis and isoprenoid synthesis in DNA replication and cell growth.J. Lipid Res. 251462–1468.Google Scholar
- Togari, A., Dickens, G., Kuzoya, H., and Guroff, G. (1985). The effect of fibroblast growth factor on PC12 cells.J. Neurosci. 5307–316.Google Scholar
- Volpe, J. J., Goldberg, R. I., and Bhat, N. R. (1985). Cholesterol biosynthesis and its regulation in dissociated cell cultures of fetal rat brain: Developmental changes and the role of 3-hydroxy-3-methylgultaryl coenzyme A reductase.J. Neurochem. 45536–543.Google Scholar