Spatial Distribution Of Tubulin Mutations Conferring Resistance To Antimicrotubular Compounds

  • Alexey Y. Nyporko
  • Yaroslav B. Blume
Part of the NATO Science for Peace and Security Series C: Environmental Security book series (NAPSC)

Resistance to antimicrotubular drugs results from single amino acid replacements in α- and β-tubulin subunits. Two possible mechanisms of action of these replacements are proposed based on analyses of their spatial distribution in the three-dimensional protein model. The main mechanism of action is typical for mutations that are localized in the immediate proximity of binding sites for antimicrotubular drugs. In this case, amino acid replacements can directly influence binding site spatial structure, and result in decreased protein affinity causing resistance only to compounds binding at this site. Mutations that cause multidrug resistance can have an alternative mechanism of action. Spatial distribution of these mutations does not correlate with the ligands' binding sites. One may assume that they effect global changes in the tubulin molecule (e.g., increasing or decreasing the general level of molecular oscillations). Therefore, theses mutations can determine either nonspecific resistance to a number of different microtubule depolymerising agents and, simultaneously, hypersen-sitivity to microtubule stabilizing compounds, or vice versa.


Antimicrotubular compounds tubulin resistance mutations mechanisms of action 


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  1. 1.
    J. J. Correia, Effects of antimitotic agents on tubulin-nucleotide interactions,Pharm. Ther. 52, 127–147 (1991).CrossRefGoogle Scholar
  2. 2.
    J. J. Correa and S. Lobert, Physiochemical aspects of tubulin-interacting antimitotic drugs,Curr. Pharmaceut. Design7, 1213–1223 (2001).CrossRefGoogle Scholar
  3. 3.
    C. Dumontet, Mechanisms of action and resistance to tubulin-binding agents,Expert Opin. Investig. Drugs9, 779–788 (2000).PubMedCrossRefGoogle Scholar
  4. 4.
    W. V. Baird, Y. B. Blume, and S. Wick, in:Plant microtubules: potential for biotechnology, edited by P. Nick (Springer Verlag, Berlin, 2000), pp. 159–191.Google Scholar
  5. 5.
    G. J. McKay and L. R. Cooke, A PCR-based method to characterise and identify benzimidazole resistance in Helminthosporium solani, FEMS Microbiol. Lett. 152, 371– 378 (1997).PubMedCrossRefGoogle Scholar
  6. 6.
    L. Elard and J. F. Humbert, Importance of the mutation of amino acid 200 of the isotype 1 beta-tubulin gene in the benzimidazole resistance of the small-ruminant parasite Teladorsagia circumcincta, Parasitol. Res.85, 452–456 (1999).PubMedCrossRefGoogle Scholar
  7. 7.
    D. H. Young and V. Lewandowski, Covalent binding of the benzamide RH-4032 to tubulin in suspension-cultured tobacco cells and its application in cell-based competitive-binding assay,Plant Physiol.124, 115–124 (2000).PubMedCrossRefGoogle Scholar
  8. 8.
    N. M. Strashnyuk and Y. B. Blume, Obtaining of mutants on microtubular protein genes,Cytol. Cenetic27, 79–96 (1993).Google Scholar
  9. 9.
    A. I. Yemets and Y. B. Blume, Resistance to herbicides with antimicrotubular activity: from natural mutants to transgenic plants,Rus. J. Plant Physiol. 46, 789–796 (1999).Google Scholar
  10. 10.
    A. Y. Nyporko, A. N. Zhivolup, and Y. B. Blume, Predicting positions of new mutations of similar features via comparative analysis of the primary structure of mutant tubulins resistant to antimicrotubular compounds,Cytol. Cenetic.37, 66–75 (2003).Google Scholar
  11. 11.
    R. G. Antony, T. R. Waldin, J. A. Ray, S. W. J. Bright, and P. J. Hussey, Herbicide resistance caused by spontaneous mutation of the cytoskeletal protein tubulin,Nature393, 260–263 (1998).CrossRefGoogle Scholar
  12. 12.
    E. Yamamoto, L. Zeng, and W. V. Baird, α-Tubulins missense mutations correlate with antimicrotubule drug resistance in Eleusine indica, Plant Cell 10, 297–308 (1998).PubMedCrossRefGoogle Scholar
  13. 13.
    S. Yin, F. Cabral, and S. Veeraraghavan, Amino acid substitutions at proline 220 of β-tubulin confer resistance to paclitaxel and colcemid,Mol. Cancer Ther.6, 2798–2806 (2007).PubMedCrossRefGoogle Scholar
  14. 14.
    Y. Wang, S. Veeraraghavan, and F. Cabral, Intra-allelic suppression of a mutation that stabilizes microtubules and confers resistance to colcemid,Biochemistry43, 8965–8973 (2004).PubMedCrossRefGoogle Scholar
  15. 15.
    F. Loganzo, M. Hari, T. Annable, X. Tan, D. B. Morilla, S. Musto, A. Zask, J. Kaplan, A. A. Minnick Jr., M. K. May, S. Ayral-Kaloustian, M. S. Poruchynsky, T. Fojo, and L. M. Greenberger, Cells resistant to HTI-286 do not overexpress P-glycoprotein but have reduced drug accumulation and a point mutation in alpha-tubulin,Mol. Cancer Ther. 10, 1319–1327 (2004).Google Scholar
  16. 16.
    M. Hari, Y. Wang, S. Veeraraghavan, and F. Cabral, Mutations in α- and β-tubulin that stabilize microtubules and confer resistance to colcemid and vinblastine,Mol. Cancer Ther. 7, 597–605 (2003).Google Scholar
  17. 17.
    M. S. Poruchynsky, J. H. Kim, E. Nogales, T. Annable, F. Loganzo, L. M. Greenberger, D. L. Sackett, and T. Fojo, Tumor cells resistant to a microtubule-depolymerizing hemiasterlin analogue, HTI-286, have mutations in alpha- or beta-tubulin and increased microtubule stability,Biochemistry43, 13944–13954 (2004).PubMedCrossRefGoogle Scholar
  18. 18.
    W. J. Blackhall, M. Drogemuller, T. Schnieder, and G. von Samson-Himmelstjerna, Expression of recombinant beta-tubulin alleles fromCylicocyclus nassatus(Cyathostominae),Parasitol. Res.99(6), 687–693 (2006).PubMedCrossRefGoogle Scholar
  19. 19.
    Y. Gokmen-Polar, D. Escuin, C. D. Walls, S. E. Soule, Y. Wang, K. L. Sanders, T. M. Lavallee, M. Wang, B. D. Guenther, P. Giannakakou, and G. W. Sledge, β-Tubulin mutations are associated with resistance to 2-methoxyestradiol in MDA-MB-435 cancer cells,Cancer Res.65, 9406–9414 (2005).PubMedCrossRefGoogle Scholar
  20. 20.
    M. W. Robinson, N. McFerran, A. Trudgett, L. Hoey, and I. Fairweather, A possible model of benzimidazole binding to β-tubulin disclosed by invoking an inter-domain movement,J. Mol. Graph. Model.23, 275–284 (2004).PubMedCrossRefGoogle Scholar
  21. 21.
    M. Ghisi, R. Kaminsky, and P. Mäser, Phenotyping and genotyping ofHaemonchus contortusisolates reveals a new putative candidate mutation for benzimidazole resistance in nematodes,Vet. Parasitol. 144, 313–320 (2007).PubMedCrossRefGoogle Scholar
  22. 22.
    M. Kavallaris, A. S. Tait, B. J. Walsh, L. He, S. B. Horwitz, M. D. Norris, and M. Haber, Multiple microtubule alterations are associated with Vinca alkaloid resistance in human leukemia cells,Cancer Res.6, 15803–5809 (2001).Google Scholar
  23. 23.
    F. Ruiz, P. Dupuis-Williams, C. Klotz, F. Forquignon, M. Bergdoll, J. Beisson, and F. Koll, Genetic evidence for interaction between eta- and beta-tubulins,Eukaryot. Cell3, 212–220 (2004).PubMedCrossRefGoogle Scholar
  24. 24.
    S. W. James, C. D. Silflow, P. Stroom, and P. A. Lefebvre, A mutation in the alpha 1-tubulin gene ofChlamydomonas reinhardtiiconfers resistance to anti-microtubule herbicides,J. Cell Sci.106, 209–218 (1993).PubMedGoogle Scholar
  25. 25.
    C. Delye, Y. Menchari, S. Michel, and H. Darmency, Molecular bases for sensitivity to tubulin-binding herbicides in green foxtail,Plant Physiol.136, 3920–3932 (2004).PubMedCrossRefGoogle Scholar
  26. 26.
    S. Tresch, P. Plath, and K. Grossmann, Herbicidal cyanoacrylates with antimicrotubule mechanism of action,Pest. Manag. Sci.61, 1052–1059 (2005).PubMedCrossRefGoogle Scholar
  27. 27.
    D. B. Lowe, G. A. Swire-Clark, L. B. McCarty, ?. Whitwell, and W. V. Baird, Biology and molecular analysis of dinitroaniline-resistantPoa annua L., Int. Turfgrass Soc. Res. J.9, 1019–1025 (2001).Google Scholar
  28. 28.
    V. D. Lee and B. Huang, Missense mutations at lysine 350 in αβ-tubulin confer altered sensitivity to microtubule inhibitors in Chlamydomonas, Plant Cell 2, 1051–1057 (1990).PubMedCrossRefGoogle Scholar
  29. 29.
    M. J. Schilber and B. Huang, The colR4 and colR15 αβ-tubulin mutations inChlamydomonas reinhardtiiconfer altered sensitivities to microtubule inhibitors and herbicides by enhancing microtubule stability,J. Cell Biol.113, 605–614 (1991).CrossRefGoogle Scholar
  30. 30.
    N. S. Morissette, A. Mitra, D. Sept, and L. D. Sibley Dinitroanilines bind α-tubulin to disrupt microtubules,Mol. Biol. Cell15, 1960–1968 (2004).CrossRefGoogle Scholar
  31. 31.
    M. Takahashi, S. Matsumoto, S. Iwasaki, and I. Yahara, Molecular basis for determining the sensitivity of eukaryotes to the antimitotic drug rhizoxin,Mol. Gen. Genet. 222, 169– 175 (1990).PubMedCrossRefGoogle Scholar
  32. 32.
    G. Zou, S. H. Ying, Z. C. Shen, and M. G. Feng, Multi-sited mutations of β-tubulin are involved in benzimidazole resistance and thermotolerance of fungal biocontrol agentBeauveria bassiana, Environ. Microbiol. 8, 2096–2105 (2006).PubMedCrossRefGoogle Scholar
  33. 33.
    M. K. Jung and B. R. Oakley, Identification of an amino acid substitution in the benA, β-tubulin gene ofAspergillus nidulansthat confers thiabendazole resistance and benomyl supersensitivity,Cell Motil. Cytoskeleton17, 87–94 (1990).PubMedCrossRefGoogle Scholar
  34. 34.
    M. K. Jung, I. B. Wilder, and B. R. Oakley, Amino acid alterations in the benA (β-tubulin) gene ofAspergillus nidulansthat confer benomyl resistance,Cell Motil. Cytoskeleton22, 170–174 (1992).PubMedCrossRefGoogle Scholar
  35. 35.
    M. Fujimura, T. Kamakura, H. Inoue, and I. Yamaguchi, Amino acid alterations in the β-tubulin gene ofNeurospora crassathat confer resistance to carbendazim and diethofencar,Curr. Genet.25, 418–422 (1994).PubMedCrossRefGoogle Scholar
  36. 36.
    K. Borck and H. D. Braymer, The genetic analysis of resistance to benomyl inNeurospora crassa. J. Gen. Microbiol. 85, 51–56 (1974).PubMedGoogle Scholar
  37. 37.
    M. J. Orbach, E. B. Porro, and C. Yanofsky, Cloning and characterization of the gene for β-tubulin from ? benomyl-resistant mutant ofNeurospora crassaand its use as ? dominant selectable marker,Mol. Cell Biol.6, 2452–2461 (1986).PubMedGoogle Scholar
  38. 38.
    S. Y. Park, O. J. Jung, Y. R. Chung, and C. W. Lee, Isolation and characterization of ? benomyl-resistant form of β-tubulin-encoding gene from the phytopathogenic fungusBotryotinia fuckeliana, Mol. Cells 7, 104–109 (1997).PubMedGoogle Scholar
  39. 39.
    Z. Ma, M. A. Yoshimura, and T. J. Michailides, Identification and characterization of benzimidazole resistance inMonilinia fructicolafrom stone fruit orchards in California,Appl. Environ. Microbiol. 69, 7145–7152 (2003).PubMedCrossRefGoogle Scholar
  40. 40.
    Y. Luo, Z. Ma, and T. J. Michailides, Quantification of allele E198A in beta-tubulin conferring benzimidazole resistance inMonilinia fructicolausing real-time PCR,Pest Manag. Sci.63, 1178–1184 (2007).PubMedCrossRefGoogle Scholar
  41. 41.
    T. L. Buhr and M. B. Dickman, Isolation, characterization, and expression of a second beta-tubulin-encoding genefrom Colletotrichum gloeosporioidesf. sp. aeschynomene,Appl. Environ. Microbiol. 60, 4155–4159 (1994).PubMedGoogle Scholar
  42. 42.
    P. Leroux, R. Fritz, D. Debieu, C. Albertini, C. Lanen, J. Bach, M. Gredt, and F. Chapeland, Mechanisms of resistance to fungicides in field strains ofBotrytis cinerea, Pest. Manag. Sci. 58, 876–888 (2002).PubMedCrossRefGoogle Scholar
  43. 43.
    Z. Ma, M. A. Yoshimura, B. A. Holtz, and T. J. Michailides, Characterization and PCR-based detection of benzimidazole-resistant isolates ofMonilinia laxain California,Pest Manag. Sci.61, 449–457 (2005).PubMedCrossRefGoogle Scholar
  44. 44.
    J. H. Thomas, N. F. Neff, and D. Botstein, Isolation and characterization of mutations in the tubulin gene ofSaccharomyces cerevisiae, Genetics 112, 715–734 (1985).Google Scholar
  45. 45.
    M. Hari, F. Loganzo, T. Annable, X. Tan, S. Musto, D. B. Morilla, J. H. Nettles, J. P. Snyder, and L. M. Greenberger, Paclitaxel-resistant cells have a mutation in the paclitaxel-binding region of beta-tubulin (Asp26Glu) and less stable microtubules.Mol. Cancer Ther. 5, 270–278 (2006).PubMedCrossRefGoogle Scholar
  46. 46.
    Y. Wang, S. Yin, K. Blade, G. Cooper, D. R. Menick, and F. Cabral, Mutations at leucine 215 of β-tubulin affect paclitaxel sensitivity by two distinct mechanisms,Biochemistry45, 185–194 (2006).PubMedCrossRefGoogle Scholar
  47. 47.
    C. P. Yang, P. Verdier-Pinard, F. Wang, E. Lippaine-Horvath, L. He, D. Li, G. Hofle, I. Ojima, G. A. Orr, and S. B. Horwitz, A highly epothilone B-resistant A549 cell line with mutations in tubulin that confer drug dependence,Mol. Cancer Ther. 4, 987–995 (2005).PubMedCrossRefGoogle Scholar
  48. 48.
    L. He, C.P. Yang, S. B. Horwitz, Mutations in beta-tubulin map to domains involved in regulation of microtubule stability in epothilone-resistant cell lines,Mol. Cancer Ther. 1, 3–10 (2001).PubMedGoogle Scholar
  49. 49.
    K. M. Wiesen, S. Xia, C. P. Yang, and S. B. Horwitz, Wild-type class I beta-tubulin sensitizes Taxol-resistant breast adenocarcinoma cells harboring a β-tubulin mutation,Cancer Lett. 257, 227–235 (2007).PubMedCrossRefGoogle Scholar
  50. 50.
    M. Gonsales-Garay, L. Chang, K. Blade, D. Menick, and F. Cabral, A beta-tubulin leucine involved in microtubule assembly and pacitaxel resistance,J. Biol. Chem.274, 23875–23882 (1999).CrossRefGoogle Scholar
  51. 51.
    N. M. Verrills, C. L. Flemming, M. Liu, M. T. Ivery, G. S. Cobon, M. D. Norris, M. Haber, and M. Kavallaris, Microtubule alterations and mutations induced by desoxyepothilone B: implications for drug-target interactions,Chem Biol. 10, 597–607 (2003).PubMedCrossRefGoogle Scholar
  52. 52.
    P. Giannakakou, D. L. Sackett, Y. K. Kang, Z. Zhan, J. T. Buters, T. Fojo, and M. S. Poruchynsky, Paclitaxel-resistant human ovarian cancer cells have mutant beta-tubulins that exhibit impaired paclitaxel-driven polymerization,J. Biol. Chem.272, 17118–17125 (1997).PubMedCrossRefGoogle Scholar
  53. 53.
    P. Giannakakou, R. Gussio, E. Nogales, K. H. Downing, D. Zaharevitz, B. Bollbuck, G. Poy, D. Sackett, K. C. Nicolaou, and T. Fojo, A common pharmacophore for epothilone and taxanes: molecular basis for drug resistance conferred by tubulin mutations in human cancer cells,Proc. Natl. Acad. Sci. USA97, 2904–2909 (2000).PubMedCrossRefGoogle Scholar
  54. 54.
    S. Hasegawa, Y. Miyoshi, C. Egawa, M. Ishitobi, Y. Tamaki, M. Monden, and S. Noguchi, Mutational analysis of the class I beta-tubulin gene in human breast cancer,Int. J. Cancer101, 46–51 (2002).PubMedCrossRefGoogle Scholar
  55. 55.
    X. H. Hua, D. Genini, R. Gussio, R. Tawatao, H. Shih, T. J. Kipps, D. A. Carson, and L. M. Leoni, Biochemical genetic analysis of indanocine resistance in human leukemia,Cancer Res.61, 7248–7254 (2001).PubMedGoogle Scholar
  56. 56.
    L. A. Martello, P. Verdier-Pinard, H. J. Shen, L. He, K. Torres, G. A. Orr, and S. B. Horwitz, Elevated levels of microtubule destabilizing factors in a Taxol-resistant/dependent A549 cell line with an α-tubulin mutation,Cancer Res.63, 1207– 1213 (2003).PubMedGoogle Scholar
  57. 57.
    A. Bairoch and R. Apweiler, The SWISS-PROT protein sequence data bank and its supplement TrEMBL in 2000,Nucleic Acids Res.28, 45–48 (2000).PubMedCrossRefGoogle Scholar
  58. 58.
    J. D. Thompson, T. J. Gibson, and F. Plewniak, The ClustalX windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools,Nucleic Acids Res.24, 4876–4882 (1997).CrossRefGoogle Scholar
  59. 59.
    R. Ravelli, B. Gigant, P. Curmi, I. Jourdain, S. Lachkar, A. Sobel, and M. Knosso, Insight into tubulin regulation from a complex with colchicines and a stathmin-like domain,Nature428, 198–202 (2004).PubMedCrossRefGoogle Scholar
  60. 60.
    B. Gigant, C. Wang, R. B. Ravelli, F. Roussi, M. O. Steinmetz, P. A. Curmi, A. Sobel, and M. Knossow, Structural basis for the regulation of tubulin by vinblastine,Nature435, 519–522 (2005).PubMedCrossRefGoogle Scholar
  61. 61.
    A. Y. Nyporko, A. I. Yemets, L. A. Klimkina, and Y. B. Blume, Sensitivity ofEleusine indicacallus to trifluralin and amiprophosmethyl in correlation with the binding of these compounds to tubulin,Russ. J. Plant Physiol. 49, 413–418 (2002).CrossRefGoogle Scholar
  62. 62.
    Y. B. Blume, A. Y. Nyporko, A. I. Yemets, and W. V. Baird, Structural modelling of the interaction of plant α-tubulin with dinitroaniline and phosphoroamidate herbicides,Cell Biol. Int.27, 171–174 (2003).PubMedCrossRefGoogle Scholar
  63. 63.
    A. Mitra and D. Sept, Binding and interaction of dinitroanilines with apicomplexan and kinetoplastid α-tubulin,J. Med. Chem.49, 5226–5231 (2006).PubMedCrossRefGoogle Scholar
  64. 64.
    A. Yu. Nyporko, A. I. Yemets, and Y. B. Blume, Protozoan and plants tubulins as specific targets for dinitroanilines and phosphoroamidates: common structural features and interactive sites.Abs. of 4-th ISGO International Conference on Structural Genomics(ISGO-2006), October 22–26, Beijing, China, pp. 257–259 (2006).Google Scholar
  65. 65.
    J. P. Snyder, J. H. Nettles, B. Cornett, K. H. Downing, and E. Nogales, The binding conformation of taxol in β-tubulin: a model based on electron crystallographic density,Proc. Natl. Acad. Sci. USA98, 5312–53126 (2001).PubMedCrossRefGoogle Scholar
  66. 66.
    J. H. Nettles, H. Li, B. Cornett, J. M. Krahn, J. P. Snyder, and K.H. Downing, The binding mode of epothilone A on αβ-tubulin by electron crystallography,Science305, 866–869 (2004).PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science + Business Media B.V. 2008

Authors and Affiliations

  • Alexey Y. Nyporko
    • 1
  • Yaroslav B. Blume
    • 1
  1. 1.Institute of Cell Biology and Genetic EngineeringNational Academy of Science of UkraineKievUkraine

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