β class II tubulin predominates in normal and tumor breast tissues

Breast Cancer Research

Sharon Lobert³  John J Correia²  Anthony Frankfurter⁴  Hamed A Benghuzzi¹  Michelle A Tucci²  Nancy Stubbs Thomas³  Jennifer A Davis²  Laree Hiser³  James H Dozier² 

[1] School of Health Related Professions, University of Mississippi Medical Center, Jackson, Mississippi, USA;School of Medicine, University of Mississippi Medical Center, Jackson, Mississippi, USA;School of Nursing, University of Mississippi Medical Center, Jackson, Mississippi, USA;Department of Biology, University of Virginia, Charlottesville, Virginia, USA

关键词: tubulin isotypes;    microtubules;    breast cancer;    antimitotic agents;   

Others  :  1118794
DOI  :  10.1186/bcr631

 received in 2002-07-01, accepted in 2003-07-07,  发布年份 2003

【 摘 要 】

Background

Antimitotic chemotherapeutic agents target tubulin, the major protein in mitotic spindles. Tubulin isotype composition is thought to be both diagnostic of tumor progression and a determinant of the cellular response to chemotherapy. This implies that there is a difference in isotype composition between normal and tumor tissues.

Methods

To determine whether such a difference occurs in breast tissues, total tubulin was fractionated from lysates of paired normal and tumor breast tissues, and the amounts of β-tubulin classes I + IV, II, and III were measured by competitive enzyme-linked immunosorbent assay (ELISA). Only primary tumor tissues, before chemotherapy, were examined. Her2/neu protein amplification occurs in about 30% of breast tumors and is considered a marker for poor prognosis. To gain insight into whether tubulin isotype levels might be correlated with prognosis, ELISAs were used to quantify Her2/neu protein levels in these tissues.

Results

β-Tubulin isotype distributions in normal and tumor breast tissues were similar. The most abundant β-tubulin isotypes in these tissues were β-tubulin classes II and I + IV. Her2/neu levels in tumor tissues were 5–30-fold those in normal tissues, although there was no correlation between the Her2/neu biomarker and tubulin isotype levels.

Conclusion

These results suggest that tubulin isotype levels, alone or in combination with Her2/neu protein levels, might not be diagnostic of tumorigenesis in breast cancer. However, the presence of a broad distribution of these tubulin isotypes (for example, 40–75% β-tubulin class II) in breast tissue, in conjunction with other factors, might still be relevant to disease progression and cellular response to antimitotic drugs.

【 授权许可】

   
2003 Dozier et al., licensee BioMed Central Ltd. This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original URL.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

Figure 8.

Figure 9.

Figure 10.

【 参考文献 】

• [1]Kennedy MJ, Donehower RC, Rowinsky EK: Treatment of metastatic breast cancer with combination paclitaxel/cyclophosphamide. Semin Oncol 1995, 22(4 Suppl 8):23-27.
• [2]Davidson NG: Single-agent paclitaxel at first relapse following adjuvant chemotherapy for breast cancer. Semin Oncol 1995, 22(Suppl 14):2-6.
• [3]Jordan MA, Thrower D, Wilson L: Mechanism of inhibition of cell proliferation by vinca alkaloids. Cancer Res 1991, 51:2212-2222.
• [4]Dhamodharan R, Jordan MA, Thrower D, Wilson L, Wadsworth P: Vinblastine suppresses dynamics of individual microtubules in living interphase cells. Mol Biol Cell 1995, 6:1215-1229.
• [5]Toso RJ, Jordan MA, Farrell KW, Matsumoto B, Wilson L: Kinetic stabilization of microtubule dynamic instability in vitro by vinblastine. Biochemistry 1993, 32:1285-1293.
• [6]Panda D, Jordan MA, Chu KC, Wilson L: Differential effects of vinblastine on polymerization and dynamics at opposite microtubule ends. J Biol Chem 1996, 271:29807-29812.
• [7]Yvon AMC, Wadsworth P, Jordan MA: Taxol suppresses dynamics of individual microtubules in living human tumor cells. Mol Biol Cell 1999, 10:947-959.
• [8]Derry WB, Wilson L, Jordan MA: Substoichiometric binding of taxol suppresses microtubule dynamics. Biochemistry 1995, 34:2203-2211.
• [9]Parness J, Horwitz SB: Taxol binds to polymerized tubulin in vitro. J Cell Biol 1981, 91:479-487.
• [10]Sullivan KF, Cleveland DW: Identification of conserved isotype-defining variable region sequences for four vertebrate β-tubulin polypeptide classes. Proc Natl Acad Sci USA 1986, 83:4327-4331.
• [11]Sullivan KF: Structure and utilization of tubulin isotypes. Annu Rev Cell Biol 1988, 4:687-716.
• [12]Lobert S, Frankfurter A, Correia JJ: Energetics of vinca alkaloid interactions with tubulin isotypes: implications for drug efficacy and toxicity. Cell Motil Cytoskel 1998, 39:107-121.
• [13]Lobert S, Frankfurter A, Correia JJ: Binding of vinblastine to phosphocellulose-purified and αβ-class III tubulin: the role of nucleotides and β-tubulin isotypes. Biochemistry 1995, 34:8050-8060.
• [14]Lee MK, Tuttle JB, Rebhun LI, Cleveland DW, Frankfurter A: The expression and posttranslational modification of neuron-specific β-tubulin isotype during chick embryogenesis. Cell Motil Cytoskel 1990, 17:118-132.
• [15]Lewis SA, Gu W, Cowan NJ: Free intermingling of mammalian β-tubulin isotypes among functionally distinct microtubules. Cell 1987, 49:539-548.
• [16]Lopata MA, Cleveland DW: In vivo microtubules are copolymers of available β-tubulin isotypes: localization of each of six vertebrate β-tubulin isotypes using polyclonal antibodies elicited by synthetic peptide antigens. J Cell Biol 1987, 105:1707-1720.
• [17]Luduena RF: Are tubulin isotypes functionally significant? Mol Biol Cell 1993, 4:445-457.
• [18]Raff EC: The role of multiple tubulin isoforms in cellular microtubule function. In In Microtubules. Edited by Hyams JS, Lloyd CW. New York: Wiley-Liss Inc; 1994:85-109.
• [19]Lewis SA, Gwo-Shu Lee M, Cowan NJ: Five mouse tubulin isotypes and their regulated expression during development. J Cell Biol 1985, 101:852-861.
• [20]Hoffman PN, Cleveland DW: Neurofilament and tubulin expression recapitulates the developmental program during axonal regeneration: induction of a specific β-tubulin isotype. Proc Natl Acad Sci USA 1988, 85:4530-4533.
• [21]Banerjee A, D'Hoore A, Engelborghs Y: Interaction of desacetamidocolchicine a fast binding analogue of colchicine with isotypically pure tubulin dimers αβ II, αβ III and αβ IV. J Biol Chem 1994, 269:10324-10329.
• [22]Lu Q, Luduena RF: Removal of β III isotype enhances taxol induced microtubule assembly. Cell Struct Funct 1993, 18:173-182.
• [23]Giannakakou P, Sackett DL, Kang Y-K, Zhan A, Buters JTM, Fojo T, Poruchynsky MS: Paclitaxel-resistant human ovarian cancer cells have mutant β-tubulins that exhibit impaired paclitaxel-driven polymerization. J Biol Chem 1997, 272:17118-17125.
• [24]Derry WB, Wilson L, Khan IA, Luduena RF, Jordan MA: Taxol differentially modulates the dynamics of microtubules assembled from unfractionated and purified β-tubulin isotypes. Biochemistry 1997, 36:3554-3562.
• [25]Laing N, Dahllof B, Hartley-Asp B, Ranganathan S, Tew K: Interaction of estramustine with tubulin isotypes. Biochemistry 1997, 36:871-878.
• [26]Falconer MM, Echeverri CJ, Brown DL: Differential sorting of β-tubulin into colchicine stable microtubules during neuronal and muscle differentiation in embryonal carcinoma cells. Cell Motil Cytoskel 1992, 21:313-325.
• [27]Haber M, Burkhart CA, Regl DL, Madafiglio J, Norris MD, Horwitz SB: Altered expression of mβ2 the class II β-tubulin isotype in murine J774 2 cell line with a high level of taxol resistance. J Biol Chem 1995, 271:31269-31275.
• [28]Kavallaris M, Kuo DY-S, Burkhart CA, Regl DL, Norris MD, Haber M, Horwitz SB: Taxol-resistant epithelial ovarian tumors are associated with altered expression of specific β-tubulin isotypes. J Clin Invest 1997, 100:1282-1293.
• [29]Ranganthan S, Benetatos CA, Colarusso PJ, Dexter DW, Hudes GR: Altered β-tubulin isotype expression in paclitaxel-resistant human prostate carcinoma cells. Br J Cancer 1998, 77:562-566.
• [30]Monzo M, Rosell R, Sanchez JJ, Lee JS, O'Brate A, Gonzalez-Larriba JL, Alberola V, Lorenzo JC, Nunez L, Ro JY, Martin C: Paclitaxel resistance in non-small-cell lung cancer associated with beta-tubulin gene mutations. J Clin Oncol 1999, 17:1786-1793.
• [31]Sangrajrang S, Denoulet P, Laing NM, Tatoud R, Millot G, Calvo F, Tew KD, Fellous A: Association of estramustine resistance in human prostatic carcinoma cells with modified patterns of tubulin expression. Biochem Pharmacol 1998, 55:325-331.
• [32]Blade K, Menick DR, Cabral F: Overexpression of class I, II or IVb β-tubulin isotypes in CHO cells is insufficient to confer resistance to paclitaxel. J Cell Sci 1999, 112:2213-2221.
• [33]Gonzalez-Garay ML, Cabral F: Overexpression of an epitope-tagged β-tubulin in Chinese hamster ovary cells causes an increase in endogenous α-tubulin synthesis. Cell Motil Cytoskel 1995, 31:259-272.
• [34]Minotti AM, Barlow SB, Cabral F: Resistance to antimitotic drugs in Chinese hamster ovary cells correlates with changes in the level of polymerized tubulin. J Biol Chem 1991, 266:3987-3994.
• [35]Cabral F, Barlow SB: Resistance to antimitotic agents as genetic probes of microtubule structure and function. Pharmacol Ther 1991, 52:159-171.
• [36]Katsetos CD, Kontogeorgos G, Geddes JF, Herman MM, Tsimara-Papastamatiou G, Yu Y, Sakkas LI, Tsokos M, Patchefsky AS, Ehya H, Coper HS, Provencio J, Spano A, Frankfurter A: Differential distribution of the neuron-associated class III β-tubulin in neuroendocrine lung tumors. Arch Pathol Lab Med 2000, 124:535-544.
• [37]Williams RC Jr, Lee JC: Preparation of tubulin from brain. Methods Enzymol 1982, 85:376-408.
• [38]Correia JJ, Baty LT, Williams RC Jr: Mg2+ dependence of guanine nucleotide binding to tubulin. J Biol Chem 1987, 262:17278-17284.
• [39]Detrich HW III, Williams RC Jr: Reversible dissociation of the αβ dimer of tubulin from bovine brain. Biochemistry 1978, 17:3900-3907.
• [40]DeFazio A, Chiew Y-E, Sini RL, Janes PW, Sutherland RL: Expression of c-erbB receptors heregulin and oestrogen receptor in human breast cell lines. Int J Cancer 2000, 87:487-498.
• [41]Gazdar AF, Kurvari V, Virmani A, Gollahon L, Sakaguchi M, Westerfield M, Kodagoda D, Stasny V, Cunningham HT, Wistuba II, Tomlinson G, Tonk V, Ashfaq R, Leitch AM, Minna JD, Shary JW: Characterization of paired tumor and non-tumor cells established from patients with breast cancer. Int J Cancer 1998, 78:766-774.
• [42]Lacey E, Snowdon KL: Isolation of mammalian brain tubulin by amino-activated gel chromatography. J Chromatogr 1990, 525:71-84.
• [43]Towbin H, Staehlin T, Gordon J: Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 1979, 76:4350-4354.
• [44]Crowther JR: Indirect ELISA. In In Methods in Molecular Biology: ELISA Theory and Practice. Totowa, NJ: Humana Press; 1995:131-160.
• [45]Banerjee A, Roach MC, Wall KA, Lopata MA, Cleveland DW, Luduena RF: A monoclonal antibody against the type II isotype of β-tubulin. J Biol Chem 1988, 263:3029-3034.
• [46]Jaffrezou J-P, Dumontet C, Derry WB, Duran G, Chen G, Tsuchiya E, Wilson L, Jordan MA, Sikic B: Novel mechanism of resistance to paclitaxel (Taxol) in human K563 leukemia cells by combined selection with PSC 833. Oncol Res 1995, 7:517-527.
• [47]Lobert S, Tucci MA, Benghuzzi HA, Frankfurter A, Correia JJ: Tubulin isotypes in normal and tumor breast tissue. Mol Biol Cell 1998, 9(Suppl):151a.
• [48]Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL: Human breast cancer: correlation of relapse and survival with amplification of the Her-2/neu oncogene. Science 1987, 235:177-181.
• [49]Wang D, Villasante A, Lewis SA, Cowan NJ: The mammalian β-tubulin repertoire hematopoietic expression of a novel heterologous β-tubulin isotype. J Cell Biol 1986, 103:1903-1910.
• [50]Murphy DB, Wallis KT: Brain and erythrocyte microtubules from chicken contain different β-tubulin polypeptides. J Biol Chem 1983, 258:7870-7875.
• [51]Gygi SP, Rochon Y, Franza R, Aebersold R: Correlation between protein and mRNA abundance in yeast. Mol Cell Biol 1999, 19:1720-1730.
• [52]Anderson L, Seilhamer J: A comparison of selected mRNA and protein abundances in human liver. Electrophoresis 1997, 18:533-537.
• [53]Tew KD, Monks A, Barone L, Rosser D, Akerman G, Montali JA, Wheatley JB, Schmidt DE Jr: Glutathione-associated enzymes in the human cell lines of the National Cancer Institute drug screening program. Mol Pharmacol 1996, 50:49-159.
• [54]Futcher B, Latter GI, Monardo P, McLaughlin CS, Garrels JI: A sampling of the yeast proteome. Mol Cell Biol 1999, 19:7357-7368.
• [55]Cleveland DW: Autoregulated control of tubulin synthesis in animal cells. Curr Opin Cell Biol 1989, 1:10-14.
• [56]Pachter JS, Yen TJ, Cleveland DW: Autoregulation of tubulin expression is achieved through specific degradation of polysomal tubulin mRNAs. Cell 1987, 51:283-292.
• [57]Yen TJ, Machlin PS, Cleveland DW: Autoregulated instability of β-tubulin mRNAs by recognition of the nascent amino terminus of β-tubulin. Nature 1988, 334:580-585.
• [58]Gonzalez-Garay ML, Cabral F: α-Tubulin limits its own synthesis: evidence for a mechanism involving translational repression. J Cell Biol 1996, 135:1525-1534.
• [59]Verdier-Pinard P, Wang F, Martello L, Burd B, Orr GA, Horwitz SB: Analysis of tubulin isotypes and mutation from taxol-restant cells by combined isoelectrofocusing and mass spectrometry. Biochemistry 2003, 42:5349-5357.
• [60]Roach MC, Boucher VL, Walss C, Ravdin PM, Luduena RF: Preparation of a monoclonal antibody specific for class I isotype of β tubulin: the β isotypes of tubulin differ in their cellular distributions within human tissues. Cell Motil Cytoskel 1998, 39:273-285.
• [61]Miahe A, Lafanechere L, Treilleux I, Peloux N, Dumontet C, Bremond A, Panh M-H, Payan R, Wehland J, Margolis R-L, Job D: Tubulin detryrosination is a frequent occurrence in breast cancers of poor prognosis. Cancer Res 2001, 61:5024-5027.
• [62]Joshi H, Cleveland D: Differential utilization of β-tubulin isotypes in differentiating neurites. J Cell Biol 1989, 109:663-673.
• [63]Thrower D, Jordan MS, Wilson L: A quantitative solid-phase assay for tubulin. Methods Cell Biol 1993, 37:29-145.
• [64]Draberova E, Lukas Z, Ivanyi D, Viklicky V, Draber P: Expression of class III β-tubulin in normal and neoplastic human tissues. Histochem Cell Biol 1998, 109:231-239.
• [65]Kavallaris M, Burkhart CA, Horwitz SB: Antisense oligonucleotides to class III β-tubuli sensitize drug-resistant cells to taxol. Br J Cancer 1999, 80:1020-1025.
• [66]Sawada T, Cabral F: Expression and function of β-tubulin isotypes in Chinese hamster ovary cells. J Biol Chem 1989, 264:3013-3020.
• [67]Kavallaris M, Tait AS, Walsh BJ, He L, Horwitz SB, Norris MD, Haber M: Multiple microtubule alterations are associated with vinca alkaloid resistance in human leukemia cells. Cancer Res 2001, 61:5803-5809.
• [68]Hari M, Yang H, Canizales M, Zeng C, Cabral F: Expression of class III β-tubulin in CHO cells reduces microtubule stability and confers resistance to paclitaxel. Proc Am Assoc Cancer Res 2002, 43:LB139.
• [69]Gonzalez-Garay ML, Chang L, Blade K, Menick DR, Cabral F: A β-tubulin leucine cluster involved in microtubule assembly and paclitaxel resistance. J Biol Chem 1999, 274:23875-23882.
• [70]Cassimeris L: Accessory protein regulation of microtubule dynamics throughout the cell cycle. Curr Opin Cell Biol 1999, 11:134-141.