Skip to main content
Plant Physiology logoLink to Plant Physiology
. 1995 Sep;109(1):277–284. doi: 10.1104/pp.109.1.277

Changes in Protein Isoprenylation during the Growth of Suspension-Cultured Tobacco Cells.

T A Morehead 1, B J Biermann 1, D N Crowell 1, S K Randall 1
PMCID: PMC157586  PMID: 12228594

Abstract

Isoprenylation facilitates the association of proteins with intracellular membranes and/or other proteins. In mammalian and yeast cells, isoprenylated proteins are involved in signal transduction, cell division, organization of the cytoskeleton, and vesicular transport. Recently, protein isoprenylation has been demonstrated in higher plants, but little is currently known about the functions of isoprenylated plant proteins. We report that inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (lovastatin) or prenyl:protein transferases (perilly alcohol) severely impair the growth of cultured tobacco (Nicotiana tabacum) cells but only when added within the first 2 d following transfer to fresh medium, before any increase in culture volume is detectable. This "window" of sensitivity to inhibitors of protein isoprenylation correlates temporally with an increase in [14C]mevalonate incorporation into tobacco cell proteins in vitro. We have also observed a marked increase in farnesyl:protein transferase activity at this early time in the growth of tobacco cultures. In contrast, type I geranylgeranyl:protein transferase activity does not change significantly during culture growth. Although these events coincide with the replication of DNA, I [mu]M lovastatin-treated cells are capable of DNA synthesis, suggesting that lovastatin-induced cell growth arrest is not due to inhibition of DNA replication. Together, these data support the hypothesis that protein isoprenylation is necessary for the early stages of growth of tobacco cultures.

Full Text

The Full Text of this article is available as a PDF (1.8 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alberts A. W., Chen J., Kuron G., Hunt V., Huff J., Hoffman C., Rothrock J., Lopez M., Joshua H., Harris E. Mevinolin: a highly potent competitive inhibitor of hydroxymethylglutaryl-coenzyme A reductase and a cholesterol-lowering agent. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3957–3961. doi: 10.1073/pnas.77.7.3957. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Balch W. E. Small GTP-binding proteins in vesicular transport. Trends Biochem Sci. 1990 Dec;15(12):473–477. doi: 10.1016/0968-0004(90)90301-q. [DOI] [PubMed] [Google Scholar]
  3. Beranger F., Cadwallader K., Porfiri E., Powers S., Evans T., de Gunzburg J., Hancock J. F. Determination of structural requirements for the interaction of Rab6 with RabGDI and Rab geranylgeranyltransferase. J Biol Chem. 1994 May 6;269(18):13637–13643. [PubMed] [Google Scholar]
  4. Biermann B. J., Morehead T. A., Tate S. E., Price J. R., Randall S. K., Crowell D. N. Novel isoprenylated proteins identified by an expression library screen. J Biol Chem. 1994 Oct 14;269(41):25251–25254. [PubMed] [Google Scholar]
  5. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  6. Brown M. S., Goldstein J. L. Multivalent feedback regulation of HMG CoA reductase, a control mechanism coordinating isoprenoid synthesis and cell growth. J Lipid Res. 1980 Jul;21(5):505–517. [PubMed] [Google Scholar]
  7. Casey P. J., Solski P. A., Der C. J., Buss J. E. p21ras is modified by a farnesyl isoprenoid. Proc Natl Acad Sci U S A. 1989 Nov;86(21):8323–8327. doi: 10.1073/pnas.86.21.8323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clarke S., Vogel J. P., Deschenes R. J., Stock J. Posttranslational modification of the Ha-ras oncogene protein: evidence for a third class of protein carboxyl methyltransferases. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4643–4647. doi: 10.1073/pnas.85.13.4643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Crowell D. N., Salaz M. S. Inhibition of growth of cultured tobacco cells at low concentrations of lovastatin is reversed by cytokinin. Plant Physiol. 1992 Dec;100(4):2090–2095. doi: 10.1104/pp.100.4.2090. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Didsbury J. R., Uhing R. J., Snyderman R. Isoprenylation of the low molecular mass GTP-binding proteins rac 1 and rac 2: possible role in membrane localization. Biochem Biophys Res Commun. 1990 Sep 14;171(2):804–812. doi: 10.1016/0006-291x(90)91217-g. [DOI] [PubMed] [Google Scholar]
  11. Fairley-Grenot K., Assmann S. M. Evidence for G-Protein Regulation of Inward K+ Channel Current in Guard Cells of Fava Bean. Plant Cell. 1991 Sep;3(9):1037–1044. doi: 10.1105/tpc.3.9.1037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Farnsworth C. C., Gelb M. H., Glomset J. A. Identification of geranylgeranyl-modified proteins in HeLa cells. Science. 1990 Jan 19;247(4940):320–322. doi: 10.1126/science.2296721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fukada Y., Takao T., Ohguro H., Yoshizawa T., Akino T., Shimonishi Y. Farnesylated gamma-subunit of photoreceptor G protein indispensable for GTP-binding. Nature. 1990 Aug 16;346(6285):658–660. doi: 10.1038/346658a0. [DOI] [PubMed] [Google Scholar]
  14. Gelb M. H., Tamanoi F., Yokoyama K., Ghomashchi F., Esson K., Gould M. N. The inhibition of protein prenyltransferases by oxygenated metabolites of limonene and perillyl alcohol. Cancer Lett. 1995 May 8;91(2):169–175. doi: 10.1016/0304-3835(95)03747-k. [DOI] [PubMed] [Google Scholar]
  15. Glomset J. A., Gelb M. H., Farnsworth C. C. Prenyl proteins in eukaryotic cells: a new type of membrane anchor. Trends Biochem Sci. 1990 Apr;15(4):139–142. doi: 10.1016/0968-0004(90)90213-u. [DOI] [PubMed] [Google Scholar]
  16. Goldstein J. L., Brown M. S. Regulation of the mevalonate pathway. Nature. 1990 Feb 1;343(6257):425–430. doi: 10.1038/343425a0. [DOI] [PubMed] [Google Scholar]
  17. Hancock J. F., Magee A. I., Childs J. E., Marshall C. J. All ras proteins are polyisoprenylated but only some are palmitoylated. Cell. 1989 Jun 30;57(7):1167–1177. doi: 10.1016/0092-8674(89)90054-8. [DOI] [PubMed] [Google Scholar]
  18. Huneeus V. Q., Wiley M. H., Siperstein M. D. Isopentenyladenine as a mediator of mevalonate-regulated DNA replication. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5842–5846. doi: 10.1073/pnas.77.10.5842. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Iraki N. M., Bressan R. A., Hasegawa P. M., Carpita N. C. Alteration of the physical and chemical structure of the primary cell wall of growth-limited plant cells adapted to osmotic stress. Plant Physiol. 1989 Sep;91(1):39–47. doi: 10.1104/pp.91.1.39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jackson J. H., Cochrane C. G., Bourne J. R., Solski P. A., Buss J. E., Der C. J. Farnesol modification of Kirsten-ras exon 4B protein is essential for transformation. Proc Natl Acad Sci U S A. 1990 Apr;87(8):3042–3046. doi: 10.1073/pnas.87.8.3042. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kawata M., Farnsworth C. C., Yoshida Y., Gelb M. H., Glomset J. A., Takai Y. Posttranslationally processed structure of the human platelet protein smg p21B: evidence for geranylgeranylation and carboxyl methylation of the C-terminal cysteine. Proc Natl Acad Sci U S A. 1990 Nov;87(22):8960–8964. doi: 10.1073/pnas.87.22.8960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Khosravi-Far R., Lutz R. J., Cox A. D., Conroy L., Bourne J. R., Sinensky M., Balch W. E., Buss J. E., Der C. J. Isoprenoid modification of rab proteins terminating in CC or CXC motifs. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6264–6268. doi: 10.1073/pnas.88.14.6264. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kinsella B. T., Maltese W. A. rab GTP-binding proteins with three different carboxyl-terminal cysteine motifs are modified in vivo by 20-carbon isoprenoids. J Biol Chem. 1992 Feb 25;267(6):3940–3945. [PubMed] [Google Scholar]
  24. Kohl N. E., Diehl R. E., Schaber M. D., Rands E., Soderman D. D., He B., Moores S. L., Pompliano D. L., Ferro-Novick S., Powers S. Structural homology among mammalian and Saccharomyces cerevisiae isoprenyl-protein transferases. J Biol Chem. 1991 Oct 5;266(28):18884–18888. [PubMed] [Google Scholar]
  25. Kuroda Y., Suzuki N., Kataoka T. The effect of posttranslational modifications on the interaction of Ras2 with adenylyl cyclase. Science. 1993 Jan 29;259(5095):683–686. doi: 10.1126/science.8430318. [DOI] [PubMed] [Google Scholar]
  26. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  27. Langan T. J., Volpe J. J. Cell cycle-specific requirement for mevalonate, but not for cholesterol, for DNA synthesis in glial primary cultures. J Neurochem. 1987 Aug;49(2):513–521. doi: 10.1111/j.1471-4159.1987.tb02894.x. [DOI] [PubMed] [Google Scholar]
  28. Li H., Roux S. J. Casein kinase II protein kinase is bound to lamina-matrix and phosphorylates lamin-like protein in isolated pea nuclei. Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8434–8438. doi: 10.1073/pnas.89.18.8434. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lutz R. J., Trujillo M. A., Denham K. S., Wenger L., Sinensky M. Nucleoplasmic localization of prelamin A: implications for prenylation-dependent lamin A assembly into the nuclear lamina. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):3000–3004. doi: 10.1073/pnas.89.7.3000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Maltese W. A., Erdman R. A. Characterization of isoprenoid involved in the post-translational modification of mammalian cell proteins. J Biol Chem. 1989 Oct 25;264(30):18168–18172. [PubMed] [Google Scholar]
  31. Maltese W. A. Posttranslational modification of proteins by isoprenoids in mammalian cells. FASEB J. 1990 Dec;4(15):3319–3328. doi: 10.1096/fasebj.4.15.2123808. [DOI] [PubMed] [Google Scholar]
  32. Maltese W. A., Robishaw J. D. Isoprenylation of C-terminal cysteine in a G-protein gamma subunit. J Biol Chem. 1990 Oct 25;265(30):18071–18074. [PubMed] [Google Scholar]
  33. Maltese W. A., Sheridan K. M., Repko E. M., Erdman R. A. Post-translational modification of low molecular mass GTP-binding proteins by isoprenoid. J Biol Chem. 1990 Feb 5;265(4):2148–2155. [PubMed] [Google Scholar]
  34. McNulty A. K., Saunders M. J. Purification and immunological detection of pea nuclear intermediate filaments: evidence for plant nuclear lamins. J Cell Sci. 1992 Oct;103(Pt 2):407–414. doi: 10.1242/jcs.103.2.407. [DOI] [PubMed] [Google Scholar]
  35. Moores S. L., Schaber M. D., Mosser S. D., Rands E., O'Hara M. B., Garsky V. M., Marshall M. S., Pompliano D. L., Gibbs J. B. Sequence dependence of protein isoprenylation. J Biol Chem. 1991 Aug 5;266(22):14603–14610. [PubMed] [Google Scholar]
  36. Mumby S. M., Casey P. J., Gilman A. G., Gutowski S., Sternweis P. C. G protein gamma subunits contain a 20-carbon isoprenoid. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5873–5877. doi: 10.1073/pnas.87.15.5873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Randall S. K., Marshall M. S., Crowell D. N. Protein isoprenylation in suspension-cultured tobacco cells. Plant Cell. 1993 Apr;5(4):433–442. doi: 10.1105/tpc.5.4.433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rasmussen C. D., Means A. R. Calmodulin is required for cell-cycle progression during G1 and mitosis. EMBO J. 1989 Jan;8(1):73–82. doi: 10.1002/j.1460-2075.1989.tb03350.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Reiss Y., Goldstein J. L., Seabra M. C., Casey P. J., Brown M. S. Inhibition of purified p21ras farnesyl:protein transferase by Cys-AAX tetrapeptides. Cell. 1990 Jul 13;62(1):81–88. doi: 10.1016/0092-8674(90)90242-7. [DOI] [PubMed] [Google Scholar]
  40. Ridley A. J., Paterson H. F., Johnston C. L., Diekmann D., Hall A. The small GTP-binding protein rac regulates growth factor-induced membrane ruffling. Cell. 1992 Aug 7;70(3):401–410. doi: 10.1016/0092-8674(92)90164-8. [DOI] [PubMed] [Google Scholar]
  41. Rothman J. E., Orci L. Molecular dissection of the secretory pathway. Nature. 1992 Jan 30;355(6359):409–415. doi: 10.1038/355409a0. [DOI] [PubMed] [Google Scholar]
  42. Seabra M. C., Goldstein J. L., Südhof T. C., Brown M. S. Rab geranylgeranyl transferase. A multisubunit enzyme that prenylates GTP-binding proteins terminating in Cys-X-Cys or Cys-Cys. J Biol Chem. 1992 Jul 15;267(20):14497–14503. [PubMed] [Google Scholar]
  43. Sinensky M., McLain T., Fantle K. Expression of prelamin A but not mature lamin A confers sensitivity of DNA biosynthesis to lovastatin on F9 teratocarcinoma cells. J Cell Sci. 1994 Aug;107(Pt 8):2215–2218. doi: 10.1242/jcs.107.8.2215. [DOI] [PubMed] [Google Scholar]
  44. Swiezewska E., Thelin A., Dallner G., Andersson B., Ernster L. Occurrence of prenylated proteins in plant cells. Biochem Biophys Res Commun. 1993 Apr 15;192(1):161–166. doi: 10.1006/bbrc.1993.1395. [DOI] [PubMed] [Google Scholar]
  45. Terryn N., Van Montagu M., Inzé D. GTP-binding proteins in plants. Plant Mol Biol. 1993 Apr;22(1):143–152. doi: 10.1007/BF00039002. [DOI] [PubMed] [Google Scholar]
  46. Tong C. G., Dauwalder M., Clawson G. A., Hatem C. L., Roux S. J. The major nucleoside triphosphatase in pea (Pisum sativum L.) nuclei and in rat liver nuclei share common epitopes also present in nuclear lamins. Plant Physiol. 1993 Mar;101(3):1005–1011. doi: 10.1104/pp.101.3.1005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Verma DPS., Cheon C., 3rd, Hong Z. Small GTP-Binding Proteins and Membrane Biogenesis in Plants. Plant Physiol. 1994 Sep;106(1):1–6. doi: 10.1104/pp.106.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Vorburger K., Kitten G. T., Nigg E. A. Modification of nuclear lamin proteins by a mevalonic acid derivative occurs in reticulocyte lysates and requires the cysteine residue of the C-terminal CXXM motif. EMBO J. 1989 Dec 20;8(13):4007–4013. doi: 10.1002/j.1460-2075.1989.tb08583.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Wolda S. L., Glomset J. A. Evidence for modification of lamin B by a product of mevalonic acid. J Biol Chem. 1988 May 5;263(13):5997–6000. [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES