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Mol Cell Biol. 1987 October; 7(10): 3792–3798.
PMCID: PMC368036

Domains of beta-tubulin essential for conserved functions in vivo.

Abstract

The relationship between the primary sequence of tubulins and their properties in cells was studied by gene transfection experiments. Previously, we studied a chimeric beta-tubulin formed from chicken beta-tubulin-2 sequences in the amino-terminal portion and the highly divergent Saccharomyces cerevisiae TUB2 sequences in the carboxy-terminal 25% of the molecule. In the cytoplasm of cultured animal cells, this protein incorporates into all microtubule structures and assembles with the same efficiency as endogenous tubulin. We show that the protein products of chimeric genes with an increasing proportion of yeast sequence, extending 5' of the carboxy-terminal 25%, are abnormal in two ways. First, they assemble with a significantly lower efficiency than the original chimeric protein or the endogenous tubulins. Second, they are less stable in the cytoplasm. The results suggest that the position of the yeast sequences is crucial in determining the properties of the molecule. Results of analyses of 1 deletion mutation and 10 linker insertions in the original chimeric tubulin suggest that those changes made outside the carboxyl terminus completely disrupt assembly activity, while those made in the carboxyl terminus do not.

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Selected References

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  • Bond JF, Fridovich-Keil JL, Pillus L, Mulligan RC, Solomon F. A chicken-yeast chimeric beta-tubulin protein is incorporated into mouse microtubules in vivo. Cell. 1986 Feb 14;44(3):461–468. [PubMed]
  • Breitling F, Little M. Carboxy-terminal regions on the surface of tubulin and microtubules. Epitope locations of YOL1/34, DM1A and DM1B. J Mol Biol. 1986 May 20;189(2):367–370. [PubMed]
  • Brown HR, Erickson HP. Assembly of proteolytically cleaved tubulin. Arch Biochem Biophys. 1983 Jan;220(1):46–51. [PubMed]
  • Cleveland DW, Sullivan KF. Molecular biology and genetics of tubulin. Annu Rev Biochem. 1985;54:331–365. [PubMed]
  • Davies J, Jimenez A. A new selective agent for eukaryotic cloning vectors. Am J Trop Med Hyg. 1980 Sep;29(5 Suppl):1089–1092. [PubMed]
  • Fine RE, Taylor L. Decreased actin and tubulin synthesis in 3T3 cells after transformation by SV40 virus. Exp Cell Res. 1976 Oct 1;102(1):162–168. [PubMed]
  • Deichman GI, Kluchareva TE, Kashkina LM, Matveeva VA. Reproducibility and relation to specific and non-specific anti-tumor resistance of the tumor "sneaking through" phenomenon. Int J Cancer. 1979 Apr 15;23(4):571–584. [PubMed]
  • Guo LH, Wu R. New rapid methods for DNA sequencing based in exonuclease III digestion followed by repair synthesis. Nucleic Acids Res. 1982 Mar 25;10(6):2065–2084. [PMC free article] [PubMed]
  • Kim S, Magendantz M, Katz W, Solomon F. Development of a differentiated microtubule structure: formation of the chicken erythrocyte marginal band in vivo. J Cell Biol. 1987 Jan;104(1):51–59. [PMC free article] [PubMed]
  • Kirchner K, Mandelkow EM. Tubulin domains responsible for assembly of dimers and protofilaments. EMBO J. 1985 Sep;4(9):2397–2402. [PubMed]
  • Lewis SA, Gu W, Cowan NJ. Free intermingling of mammalian beta-tubulin isotypes among functionally distinct microtubules. Cell. 1987 May 22;49(4):539–548. [PubMed]
  • Steele JC, Jr, Tanford C, Reynolds JA. Determination of partial specific volumes for lipid-associated proteins. Methods Enzymol. 1978;48:11–23. [PubMed]
  • Ludueńa RF, Shooter EM, Wilson L. Structure of the tubulin dimer. J Biol Chem. 1977 Oct 25;252(20):7006–7014. [PubMed]
  • Maccioni RB, Serrano L, Avila J, Cann JR. Characterization and structural aspects of the enhanced assembly of tubulin after removal of its carboxyl-terminal domain. Eur J Biochem. 1986 Apr 15;156(2):375–381. [PubMed]
  • Mandelkow EM, Herrmann M, Rühl U. Tubulin domains probed by limited proteolysis and subunit-specific antibodies. J Mol Biol. 1985 Sep 20;185(2):311–327. [PubMed]
  • Mulligan RC, Berg P. Selection for animal cells that express the Escherichia coli gene coding for xanthine-guanine phosphoribosyltransferase. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2072–2076. [PubMed]
  • Neff NF, Thomas JH, Grisafi P, Botstein D. Isolation of the beta-tubulin gene from yeast and demonstration of its essential function in vivo. Cell. 1983 May;33(1):211–219. [PubMed]
  • Ng M, Privalsky ML. Structural domains of the avian erythroblastosis virus erbB protein required for fibroblast transformation: dissection by in-frame insertional mutagenesis. J Virol. 1986 May;58(2):542–553. [PMC free article] [PubMed]
  • Parker BA, Stark GR. Regulation of simian virus 40 transcription: sensitive analysis of the RNA species present early in infections by virus or viral DNA. J Virol. 1979 Aug;31(2):360–369. [PMC free article] [PubMed]
  • Sackett DL, Bhattacharyya B, Wolff J. Tubulin subunit carboxyl termini determine polymerization efficiency. J Biol Chem. 1985 Jan 10;260(1):43–45. [PubMed]
  • Sackett DL, Wolff J. Proteolysis of tubulin and the substructure of the tubulin dimer. J Biol Chem. 1986 Jul 5;261(19):9070–9076. [PubMed]
  • Schatz PJ, Georges GE, Solomon F, Botstein D. Insertions of up to 17 amino acids into a region of alpha-tubulin do not disrupt function in vivo. Mol Cell Biol. 1987 Oct;7(10):3799–3805. [PMC free article] [PubMed]
  • Schiff PB, Fant J, Horwitz SB. Promotion of microtubule assembly in vitro by taxol. Nature. 1979 Feb 22;277(5698):665–667. [PubMed]
  • Solomon F, Magendantz M, Salzman A. Identification with cellular microtubules of one of the co-assemlbing microtubule-associated proteins. Cell. 1979 Oct;18(2):431–438. [PubMed]
  • Southern PJ, Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed]
  • Theurkauf WE, Baum H, Bo J, Wensink PC. Tissue-specific and constitutive alpha-tubulin genes of Drosophila melanogaster code for structurally distinct proteins. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8477–8481. [PubMed]
  • Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. [PubMed]
  • Valenzuela P, Quiroga M, Zaldivar J, Rutter WJ, Kirschner MW, Cleveland DW. Nucleotide and corresponding amino acid sequences encoded by alpha and beta tubulin mRNAs. Nature. 1981 Feb 19;289(5799):650–655. [PubMed]
  • Zieve G, Solomon F. Proteins specifically associated with the microtubules of the mammalian mitotic spindle. Cell. 1982 Feb;28(2):233–242. [PubMed]

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