Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1995 Dec;15(12):6613–6622. doi: 10.1128/mcb.15.12.6613

Evidence that farnesyltransferase inhibitors suppress Ras transformation by interfering with Rho activity.

P F Lebowitz 1, J P Davide 1, G C Prendergast 1
PMCID: PMC230914  PMID: 8524226

Abstract

Small-molecule inhibitors of the housekeeping enzyme farnesyltransferase (FT) suppress the malignant growth of Ras-transformed cells. Previous work suggested that the activity of these compounds reflected effects on actin stress fiber regulation rather than Ras inhibition. Rho proteins regulate stress fiber formation, and one member of this family, RhoB, is farnesylated in vivo. Therefore, we tested the hypothesis that interference with RhoB was the principal basis by which the peptidomimetic FT inhibitor L-739,749 suppressed Ras transformation. The half-life of RhoB was found to be approximately 2 h, supporting the possibility that it could be functionally depleted within the 18-h period required by L-739,749 to induce reversion. Cell treatment with L-739,749 disrupted the vesicular localization of RhoB but did not effect the localization of the closely related RhoA protein. Ras-transformed Rat1 cells ectopically expressing N-myristylated forms of RhoB (Myr-rhoB), whose vesicular localization was unaffected by L-739,749, were resistant to drug treatment. The protective effect of Myr-rhoB required the integrity of the RhoB effector domain and was not due to a gain-of-function effect of myristylation on cell growth. In contrast, Rat1 cells transformed by a myristylated Ras construct remained susceptible to growth inhibition by L-739,749. We concluded that Rho is necessary for Ras transformation and that FT inhibitors suppress the transformed phenotype at least in part by direct or indirect interference with Rho, possibly with RhoB itself.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Adamson P., Marshall C. J., Hall A., Tilbrook P. A. Post-translational modifications of p21rho proteins. J Biol Chem. 1992 Oct 5;267(28):20033–20038. [PubMed] [Google Scholar]
  2. Adamson P., Paterson H. F., Hall A. Intracellular localization of the P21rho proteins. J Cell Biol. 1992 Nov;119(3):617–627. doi: 10.1083/jcb.119.3.617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Armstrong S. A., Hannah V. C., Goldstein J. L., Brown M. S. CAAX geranylgeranyl transferase transfers farnesyl as efficiently as geranylgeranyl to RhoB. J Biol Chem. 1995 Apr 7;270(14):7864–7868. doi: 10.1074/jbc.270.14.7864. [DOI] [PubMed] [Google Scholar]
  4. Bar-Sagi D., Feramisco J. R. Induction of membrane ruffling and fluid-phase pinocytosis in quiescent fibroblasts by ras proteins. Science. 1986 Sep 5;233(4768):1061–1068. doi: 10.1126/science.3090687. [DOI] [PubMed] [Google Scholar]
  5. Brown M. S., Goldstein J. L. Protein prenylation. Mad bet for Rab. Nature. 1993 Nov 4;366(6450):14–15. doi: 10.1038/366014a0. [DOI] [PubMed] [Google Scholar]
  6. Buss J. E., Solski P. A., Schaeffer J. P., MacDonald M. J., Der C. J. Activation of the cellular proto-oncogene product p21Ras by addition of a myristylation signal. Science. 1989 Mar 24;243(4898):1600–1603. doi: 10.1126/science.2648572. [DOI] [PubMed] [Google Scholar]
  7. Chen C., Okayama H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol. 1987 Aug;7(8):2745–2752. doi: 10.1128/mcb.7.8.2745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clarke S. Protein isoprenylation and methylation at carboxyl-terminal cysteine residues. Annu Rev Biochem. 1992;61:355–386. doi: 10.1146/annurev.bi.61.070192.002035. [DOI] [PubMed] [Google Scholar]
  9. Cox A. D., Der C. J. Biological assays for cellular transformation. Methods Enzymol. 1994;238:277–294. doi: 10.1016/0076-6879(94)38026-0. [DOI] [PubMed] [Google Scholar]
  10. Cox A. D., Garcia A. M., Westwick J. K., Kowalczyk J. J., Lewis M. D., Brenner D. A., Der C. J. The CAAX peptidomimetic compound B581 specifically blocks farnesylated, but not geranylgeranylated or myristylated, oncogenic ras signaling and transformation. J Biol Chem. 1994 Jul 29;269(30):19203–19206. [PubMed] [Google Scholar]
  11. Dartsch P. C., Ritter M., Häussinger D., Lang F. Cytoskeletal reorganization in NIH 3T3 fibroblasts expressing the ras oncogene. Eur J Cell Biol. 1994 Apr;63(2):316–325. [PubMed] [Google Scholar]
  12. DeClue J. E., Vass W. C., Papageorge A. G., Lowy D. R., Willumsen B. M. Inhibition of cell growth by lovastatin is independent of ras function. Cancer Res. 1991 Jan 15;51(2):712–717. [PubMed] [Google Scholar]
  13. Farnsworth C. C., Wolda S. L., Gelb M. H., Glomset J. A. Human lamin B contains a farnesylated cysteine residue. J Biol Chem. 1989 Dec 5;264(34):20422–20429. [PMC free article] [PubMed] [Google Scholar]
  14. Garcia A. M., Rowell C., Ackermann K., Kowalczyk J. J., Lewis M. D. Peptidomimetic inhibitors of Ras farnesylation and function in whole cells. J Biol Chem. 1993 Sep 5;268(25):18415–18418. [PubMed] [Google Scholar]
  15. Gibbs J. B., Oliff A., Kohl N. E. Farnesyltransferase inhibitors: Ras research yields a potential cancer therapeutic. Cell. 1994 Apr 22;77(2):175–178. doi: 10.1016/0092-8674(94)90308-5. [DOI] [PubMed] [Google Scholar]
  16. Gibbs J. B. Ras C-terminal processing enzymes--new drug targets? Cell. 1991 Apr 5;65(1):1–4. doi: 10.1016/0092-8674(91)90352-y. [DOI] [PubMed] [Google Scholar]
  17. 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]
  18. Hall A. Ras-related GTPases and the cytoskeleton. Mol Biol Cell. 1992 May;3(5):475–479. doi: 10.1091/mbc.3.5.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hancock J. F., Paterson H., Marshall C. J. A polybasic domain or palmitoylation is required in addition to the CAAX motif to localize p21ras to the plasma membrane. Cell. 1990 Oct 5;63(1):133–139. doi: 10.1016/0092-8674(90)90294-o. [DOI] [PubMed] [Google Scholar]
  20. Ingber D. E., Dike L., Hansen L., Karp S., Liley H., Maniotis A., McNamee H., Mooney D., Plopper G., Sims J. Cellular tensegrity: exploring how mechanical changes in the cytoskeleton regulate cell growth, migration, and tissue pattern during morphogenesis. Int Rev Cytol. 1994;150:173–224. doi: 10.1016/s0074-7696(08)61542-9. [DOI] [PubMed] [Google Scholar]
  21. James G. L., Goldstein J. L., Brown M. S., Rawson T. E., Somers T. C., McDowell R. S., Crowley C. W., Lucas B. K., Levinson A. D., Marsters J. C., Jr Benzodiazepine peptidomimetics: potent inhibitors of Ras farnesylation in animal cells. Science. 1993 Jun 25;260(5116):1937–1942. doi: 10.1126/science.8316834. [DOI] [PubMed] [Google Scholar]
  22. Jähner D., Hunter T. The ras-related gene rhoB is an immediate-early gene inducible by v-Fps, epidermal growth factor, and platelet-derived growth factor in rat fibroblasts. Mol Cell Biol. 1991 Jul;11(7):3682–3690. doi: 10.1128/mcb.11.7.3682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Katayama M., Kawata M., Yoshida Y., Horiuchi H., Yamamoto T., Matsuura Y., Takai Y. The posttranslationally modified C-terminal structure of bovine aortic smooth muscle rhoA p21. J Biol Chem. 1991 Jul 5;266(19):12639–12645. [PubMed] [Google Scholar]
  24. Kato K., Cox A. D., Hisaka M. M., Graham S. M., Buss J. E., Der C. J. Isoprenoid addition to Ras protein is the critical modification for its membrane association and transforming activity. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6403–6407. doi: 10.1073/pnas.89.14.6403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Khosravi-Far R., Solski P. A., Clark G. J., Kinch M. S., Der C. J. Activation of Rac1, RhoA, and mitogen-activated protein kinases is required for Ras transformation. Mol Cell Biol. 1995 Nov;15(11):6443–6453. doi: 10.1128/mcb.15.11.6443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kohl N. E., Mosser S. D., deSolms S. J., Giuliani E. A., Pompliano D. L., Graham S. L., Smith R. L., Scolnick E. M., Oliff A., Gibbs J. B. Selective inhibition of ras-dependent transformation by a farnesyltransferase inhibitor. Science. 1993 Jun 25;260(5116):1934–1937. doi: 10.1126/science.8316833. [DOI] [PubMed] [Google Scholar]
  27. Kohl N. E., Omer C. A., Conner M. W., Anthony N. J., Davide J. P., deSolms S. J., Giuliani E. A., Gomez R. P., Graham S. L., Hamilton K. Inhibition of farnesyltransferase induces regression of mammary and salivary carcinomas in ras transgenic mice. Nat Med. 1995 Aug;1(8):792–797. doi: 10.1038/nm0895-792. [DOI] [PubMed] [Google Scholar]
  28. Kohl N. E., Wilson F. R., Mosser S. D., Giuliani E., deSolms S. J., Conner M. W., Anthony N. J., Holtz W. J., Gomez R. P., Lee T. J. Protein farnesyltransferase inhibitors block the growth of ras-dependent tumors in nude mice. Proc Natl Acad Sci U S A. 1994 Sep 13;91(19):9141–9145. doi: 10.1073/pnas.91.19.9141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lacal P. M., Pennington C. Y., Lacal J. C. Transforming activity of ras proteins translocated to the plasma membrane by a myristoylation sequence from the src gene product. Oncogene. 1988 Jun;2(6):533–537. [PubMed] [Google Scholar]
  30. Lo S. H., Chen L. B. Focal adhesion as a signal transduction organelle. Cancer Metastasis Rev. 1994 Mar;13(1):9–24. doi: 10.1007/BF00690415. [DOI] [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. Manne V., Yan N., Carboni J. M., Tuomari A. V., Ricca C. S., Brown J. G., Andahazy M. L., Schmidt R. J., Patel D., Zahler R. Bisubstrate inhibitors of farnesyltransferase: a novel class of specific inhibitors of ras transformed cells. Oncogene. 1995 May 4;10(9):1763–1779. [PubMed] [Google Scholar]
  33. Marshall C. J. Protein prenylation: a mediator of protein-protein interactions. Science. 1993 Mar 26;259(5103):1865–1866. doi: 10.1126/science.8456312. [DOI] [PubMed] [Google Scholar]
  34. 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]
  35. Newman C. M., Magee A. I. Posttranslational processing of the ras superfamily of small GTP-binding proteins. Biochim Biophys Acta. 1993 May 25;1155(1):79–96. doi: 10.1016/0304-419x(93)90023-6. [DOI] [PubMed] [Google Scholar]
  36. Niman H. L., Houghten R. A., Walker L. E., Reisfeld R. A., Wilson I. A., Hogle J. M., Lerner R. A. Generation of protein-reactive antibodies by short peptides is an event of high frequency: implications for the structural basis of immune recognition. Proc Natl Acad Sci U S A. 1983 Aug;80(16):4949–4953. doi: 10.1073/pnas.80.16.4949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Paterson H. F., Self A. J., Garrett M. D., Just I., Aktories K., Hall A. Microinjection of recombinant p21rho induces rapid changes in cell morphology. J Cell Biol. 1990 Sep;111(3):1001–1007. doi: 10.1083/jcb.111.3.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pompliano D. L., Rands E., Schaber M. D., Mosser S. D., Anthony N. J., Gibbs J. B. Steady-state kinetic mechanism of Ras farnesyl:protein transferase. Biochemistry. 1992 Apr 21;31(15):3800–3807. doi: 10.1021/bi00130a010. [DOI] [PubMed] [Google Scholar]
  39. Prendergast G. C., Davide J. P., Kral A., Diehl R., Gibbs J. B., Omer C. A., Kohl N. E. Negative growth selection against rodent fibroblasts targeted for genetic inhibition of farnesyl transferase. Cell Growth Differ. 1993 Sep;4(9):707–713. [PubMed] [Google Scholar]
  40. Prendergast G. C., Davide J. P., deSolms S. J., Giuliani E. A., Graham S. L., Gibbs J. B., Oliff A., Kohl N. E. Farnesyltransferase inhibition causes morphological reversion of ras-transformed cells by a complex mechanism that involves regulation of the actin cytoskeleton. Mol Cell Biol. 1994 Jun;14(6):4193–4202. doi: 10.1128/mcb.14.6.4193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Prendergast G. C., Diamond L. E., Dahl D., Cole M. D. The c-myc-regulated gene mrl encodes plasminogen activator inhibitor 1. Mol Cell Biol. 1990 Mar;10(3):1265–1269. doi: 10.1128/mcb.10.3.1265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Prendergast G. C., Khosravi-Far R., Solski P. A., Kurzawa H., Lebowitz P. F., Der C. J. Critical role of Rho in cell transformation by oncogenic Ras. Oncogene. 1995 Jun 15;10(12):2289–2296. [PubMed] [Google Scholar]
  43. Prendergast G. C., Ziff E. B. Mbh 1: a novel gelsolin/severin-related protein which binds actin in vitro and exhibits nuclear localization in vivo. EMBO J. 1991 Apr;10(4):757–766. doi: 10.1002/j.1460-2075.1991.tb08007.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Qiu R. G., Chen J., Kirn D., McCormick F., Symons M. An essential role for Rac in Ras transformation. Nature. 1995 Mar 30;374(6521):457–459. doi: 10.1038/374457a0. [DOI] [PubMed] [Google Scholar]
  45. Ridley A. J., Hall A. The small GTP-binding protein rho regulates the assembly of focal adhesions and actin stress fibers in response to growth factors. Cell. 1992 Aug 7;70(3):389–399. doi: 10.1016/0092-8674(92)90163-7. [DOI] [PubMed] [Google Scholar]
  46. Ridley A. J. Membrane ruffling and signal transduction. Bioessays. 1994 May;16(5):321–327. doi: 10.1002/bies.950160506. [DOI] [PubMed] [Google Scholar]
  47. 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]
  48. Schafer W. R., Rine J. Protein prenylation: genes, enzymes, targets, and functions. Annu Rev Genet. 1992;26:209–237. doi: 10.1146/annurev.ge.26.120192.001233. [DOI] [PubMed] [Google Scholar]
  49. Schlaepfer D. D., Hanks S. K., Hunter T., van der Geer P. Integrin-mediated signal transduction linked to Ras pathway by GRB2 binding to focal adhesion kinase. Nature. 1994 Dec 22;372(6508):786–791. doi: 10.1038/372786a0. [DOI] [PubMed] [Google Scholar]
  50. Ulsh L. S., Shih T. Y. Metabolic turnover of human c-rasH p21 protein of EJ bladder carcinoma and its normal cellular and viral homologs. Mol Cell Biol. 1984 Aug;4(8):1647–1652. doi: 10.1128/mcb.4.8.1647. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wyss A., Kaech S., Ballmer-Hofer K. Myristylation of pp60c-src is not required for complex formation with polyomavirus middle-T antigen. J Virol. 1990 Oct;64(10):5163–5166. doi: 10.1128/jvi.64.10.5163-5166.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Yamamoto M., Marui N., Sakai T., Morii N., Kozaki S., Ikai K., Imamura S., Narumiya S. ADP-ribosylation of the rhoA gene product by botulinum C3 exoenzyme causes Swiss 3T3 cells to accumulate in the G1 phase of the cell cycle. Oncogene. 1993 Jun;8(6):1449–1455. [PubMed] [Google Scholar]
  53. Yeramian P., Chardin P., Madaule P., Tavitian A. Nucleotide sequence of human rho cDNA clone 12. Nucleic Acids Res. 1987 Feb 25;15(4):1869–1869. doi: 10.1093/nar/15.4.1869. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES