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Stabilization and bundling of subtilisin-treated microtubules induced by microtubule associated proteins
Y. Saoudi, I. Paintrand, L. Multigner, D. Job
Journal of Cell Science 1995 108: 357-367;

Summary

The acidic carboxy-terminal regions of alpha- and beta-tubulin subunits are currently thought to be centrally involved in microtubule stability and in microtubule association with a variety of proteins (MAPs) such as MAP2 and tau proteins. Here, pure tubulin microtubules were exposed to subtilisin to produce polymers composed of cleaved tubulin subunits lacking carboxy termini. Polymer exposure to subtilisin was achieved in buffer conditions compatible with further tests of microtubule stability. Microtubules composed of normal alpha-tubulin and cleaved beta-tubulin were indistinguishable from control microtubules with regard to resistance to dilution-induced disassembly, to cold temperature-induced disassembly and to Ca(2+)-induced disassembly. Microtubules composed of cleaved alpha- and beta-tubulins showed normal sensitivity to dilution-induced disassembly and to low temperature-induced disassembly, but marked resistance to Ca(2+)-induced disassembly. Polymers composed of normal alpha-tubulin and cleaved beta-tubulin or of cleaved alpha- and beta-tubulins were stabilized in the presence of added MAP2, myelin basic protein and histone H1. Cleavage of tubulin carboxy termini greatly potentiated microtubule stabilization by tau proteins. We show that this potentiation of polymer stabilization can be ascribed to tau-induced microtubule bundling. In our working conditions, such bundling upon association with tau proteins occurred only in the case of microtubules composed of cleaved alpha- and beta-tubulins and triggered apparent microtubule cross-stabilization among the bundled polymers. These results, as well as immunofluorescence analysis, which directly showed interactions between subtilisin-treated microtubules and MAPs, suggest that the carboxy termini of alpha- and beta-tubulins are not primarily involved in the binding of MAPs onto microtubules. However, interactions between tubulin carboxy termini and MAPs remain possible and might be involved in the regulation of MAP-induced microtubule bundling.

REFERENCES

    1. Bhattacharyya B.,
    2. Sackett D. L. and
    3. Wolff J.
    (1985). Tubulin, hybrid dimers, and tubulin S. Stepwise charge reduction and polymerization. J. Biol. Chem 260, 10208–10216
    OpenUrlAbstract/FREE Full Text
    1. Breitling F. and
    2. Little M.
    (1986). Carboxy-terminal regions on the surface of tubulin and microtubules. Epitope locations of YOL1/34, DM1A and DM1B. J. Mol. Biol 189, 367–370
    OpenUrlCrossRefPubMedWeb of Science
    1. Burgess S. A.,
    2. Carter D. A.,
    3. Dover S. and
    4. Woolley D. M.
    (1991). The inner dynein arm complex: compatible images from freeze-etch and thin section methods of microscopy. J. Cell Sci 100, 319–328
    OpenUrlAbstract/FREE Full Text
    1. Cleveland D. W.,
    2. Joshi H. C. and
    3. Murphy D. B.
    (1990). Tubulin site interpretation. Nature 344, 389–.
    OpenUrlCrossRefPubMed
    1. Cross D.,
    2. Dominguez J.,
    3. Maccioni R. B. and
    4. Avila J.
    (1991). MAP-1 and MAP-2 binding sites at the C-terminus of-tubulin. Studies with synthetic tubulin peptides. Biochemistry 30, 4362–4366
    OpenUrlCrossRefPubMed
    1. Dye R. B.,
    2. Fink P. S. and
    3. Williams R. C.
    (1993). Taxol-induced flexibility of microtubules and its reversal by MAP-2 and tau. J. Biol. Chem 268, 6847–6850
    OpenUrlAbstract/FREE Full Text
    1. Grimm M.,
    2. Breitling F. and
    3. Little M.
    (1987). Localization of the epitope for the-tubulin monoclonal antibody TU.01. Biochim. Biophys. Acta 914, 83–88
    OpenUrlCrossRefPubMed
    1. Gundersen G. G.,
    2. Kalnoski M. H. and
    3. Bulinski J. C.
    (1984). Distinct populations of microtubules: tyrosinated and nontyrosinated alpha tubulin are distributed differently in vivo. Cell 38, 779–789
    OpenUrlCrossRefPubMedWeb of Science
    1. Job D.,
    2. Pabion M. and
    3. Margolis R. L.
    (1985). Generation of microtubule stability subclasses by microtubule-associated proteins: implications for the microtubule “dynamic instability” model. J. Cell Biol 101, 1680–1689
    OpenUrlAbstract/FREE Full Text
    1. Kanai Y.,
    2. Takemura R.,
    3. Oshima T.,
    4. Mori H.,
    5. Ihara Y.,
    6. Yanagisawa M.,
    7. Masaki T. and
    8. Hirokawa N.
    (1989). Expression of multiple tau isoforms and microtubule bundle formation in fibroblasts transfected with a single tau cDNA. J. Cell Biol 109, 1173–1184
    OpenUrlAbstract/FREE Full Text
    1. Laemmli U. K.
    (1970). Cleavage of structural proteins during the assembly of bacteriophage T4. Nature 227, 680–685
    OpenUrlCrossRefPubMedWeb of Science
    1. Lewis S. A.,
    2. Ivanov I. E.,
    3. Lee G. H. and
    4. Cowan N. J.
    (1989). Organization of microtubules in dendrites and axons is determined by a short hydrophobic zipper in microtubule-associated proteins MAP2 and tau. Nature 342, 498–505
    OpenUrlCrossRefPubMed
    1. Lewis S. A. and
    2. Cowan N.
    (1990). Microtubule bundling. Nature 345, 674–.
    OpenUrlCrossRefPubMed
    1. Littauer U. Z.,
    2. Giveon D.,
    3. Thierauf M.,
    4. Ginzburg I. and
    5. Ponstingl H.
    (1986). Common and distinct tubulin binding sites for microtubule-associated proteins. Proc. Nat. Acad. Sci. USA 83, 7162–7166
    OpenUrlAbstract/FREE Full Text
    1. Lobert S. and
    2. Correia J. J.
    (1992). Subtilisin cleavage of tubulin heterodimers and polymers. Arch. Biochem. Biophys 296, 152–160
    OpenUrlCrossRefPubMed
    1. Lobert S.,
    2. Hennington B. S. and
    3. Correia J. J.
    (1993). Multiple sites for subtilisin cleavage of tubulin: effects of divalent cations. Cell Motil. Cytoskel 25, 282–297
    OpenUrlCrossRefPubMed
    1. Maccioni R. B.,
    2. Serrano L.,
    3. Avila J. and
    4. Cann J. R.
    (1986). Characterization and structural aspects of the enhanced assembly of tubulin after removal of its carboxyl-terminal domain. Eur. J. Biochem 156, 375–381
    OpenUrlPubMedWeb of Science
    1. Maccioni R. B.,
    2. Rivas C. I. and
    3. Vera J. C.
    (1988). Differential interaction of synthetic peptides from the carboxyl-terminal regulatory domain of tubulin with microtubule-associated proteins. EMBO J 7, 1957–1963
    OpenUrlPubMedWeb of Science
    1. Marya P. K.,
    2. Syed Z.,
    3. Fraylich P. E. and
    4. Eagles P. A. M.
    (1994). Kinesin and tau bind to distinct sites on microtubules. J. Cell Sci 107, 339–344
    OpenUrlAbstract/FREE Full Text
    1. Matthes T.,
    2. Wolff A.,
    3. Soubiran P.,
    4. Gros F. and
    5. Dighiero G.
    (1988). Antitubulin antibodies. II. Natural autoantibodies and induced antibodies recognize different epitopes on the tubulin molecule. J. Immunol 141, 3135–3141
    OpenUrlAbstract/FREE Full Text
    1. Melki R.,
    2. Kerjan P.,
    3. Waller J. P.,
    4. Carlier M. F. and
    5. Pantaloni D.
    (1991). Interaction of microtubule-associated proteins with microtubules: yeast lysyl and valyl-tRNA synthetases and T218-235 synthetic peptide as model systems. Biochemistry 30, 11536–11545
    OpenUrlCrossRefPubMed
    1. Multigner L.,
    2. Gagnon J.,
    3. Van Dorsselaer A. and
    4. Job D.
    (1992). Stabilization of sea urchin flagellar microtubules by histone H1. Nature 360, 33–39
    OpenUrlCrossRefPubMedWeb of Science
    1. Olmsted J. B.
    (1986). Microtubule-associated proteins. Annu. Rev. Cell Biol 2, 421–457
    OpenUrlCrossRefWeb of Science
    1. Paschal B. M.,
    2. Obar R. A. and
    3. Vallee R. B.
    (1989). Interaction of brain cytoplasmic dynein and MAP2 with a common sequence at the C-terminus of tubulin. Nature 342, 569–572
    OpenUrlCrossRefPubMedWeb of Science
    1. Paturle-Lafanechere L.,
    2. Edde B.,
    3. Denoulet P.,
    4. Van Dorsselaer A.,
    5. Mazarguil H.,
    6. Le Caer J. P.,
    7. Wehland J. and
    8. Job D.
    (1991). Characterization of a major brain tubulin variant which cannot be tyrosinated. Biochemistry 30, 10523–10528
    OpenUrlCrossRefPubMedWeb of Science
    1. Paturle-Lafanechere L.,
    2. Manier M.,
    3. Trigault N.,
    4. Pirollet F.,
    5. Mazarguil H. and
    6. Job D.
    (1994). Accumulation of2-tubulin, a major tubulin variant which cannot be tyrosinated, in neuronal tissues and in stable microtubule assemblies. J. Cell Sci 107, 1529–1543
    OpenUrlAbstract/FREE Full Text
    1. Pirollet F.,
    2. Job D.,
    3. Margolis R. L. and
    4. Garel J. R.
    (1987). An oscillatory mode for microtubule assembly. EMBO J 6, 3247–3252
    OpenUrlPubMedWeb of Science
    1. Pirollet F.,
    2. Rauch C. T.,
    3. Job D. and
    4. Margolis R. L.
    (1989). Monoclonal antibody to microtubule-associated STOP protein: affinity purification of neuronal STOP activity and comparison of antigen with activity in neuronal and non-neuronal cell extracts. Biochemistry 28, 835–842
    OpenUrlCrossRefPubMed
    1. Pirollet F.,
    2. Derancourt J.,
    3. Haiech J.,
    4. Job D. and
    5. Margolis R. L.
    (1992). Ca2+-calmodulin regulated effectors of microtubule stability in bovine brain. Biochemistry 31, 8849–8855
    OpenUrlCrossRefPubMed
    1. Rodionov V. I.,
    2. Gyoeva F. K.,
    3. Kashina A. S.,
    4. Kuznetsov S. A. and
    5. Gelfand V. I.
    (1990). Microtubule-associated proteins and microtubule-based translocators have different binding sites on tubulin molecule. J. Biol. Chem 265, 5702–5707
    OpenUrlAbstract/FREE Full Text
    1. Sackett D. I.,
    2. Bhattacharyya B. and
    3. Wolff J.
    (1985). Tubulin subunit carboxyl termini determine polymerization efficiency. J. Biol. Chem 260, 43–45
    OpenUrlAbstract/FREE Full Text
    1. Scott C. W.,
    2. Klika A. B.,
    3. Lo M. M. S.,
    4. Norris T. E. and
    5. Caputo C. B.
    (1992). Tau protein induces bundling of microtubules in vitro: comparison of different tau isoforms and a tau protein fragment. J. Neurosci. Res 33, 19–29
    OpenUrlCrossRefPubMedWeb of Science
    1. Serrano L.,
    2. Avila J. and
    3. Maccioni R. B.
    (1984). Controlled proteolysis of tubulin by subtilisin: localization of the site for MAP2 interaction. Biochemistry 23, 4675–4681
    OpenUrlCrossRefPubMed
    1. Serrano L.,
    2. de la Torre J.,
    3. Maccioni R. B. and
    4. Avila J.
    (1984). Involvement of the carboxyl-terminal domain of tubulin in the regulation of its assembly. Proc. Nat. Acad. Sci. USA 81, 5989–5993
    OpenUrlAbstract/FREE Full Text
    1. Serrano L.,
    2. Montejo de Garcini E.,
    3. Hernándes M. A. and
    4. Avila J.
    (1985). Localization of the tubulin binding site for tau proteins. Eur. J. Biochem 153, 595–600
    OpenUrlPubMedWeb of Science
    1. Serrano L.,
    2. Valencia A.,
    3. Caballero R. and
    4. Avila J.
    (1986). Localization of the high affinity calcium-binding site on tubulin molecule. J. Biol. Chem 261, 7076–7081
    OpenUrlAbstract/FREE Full Text
    1. Serrano L.,
    2. Wandosell F.,
    3. de la Torre J. and
    4. Avila J.
    (1988). Effect ofspecific proteolytic cleavages on tubulin polymer formation. Biochem. J 252, 683–691
    OpenUrlAbstract/FREE Full Text
    1. Sobue K.,
    2. Fujita M.,
    3. Muramoto Y. and
    4. Kakiuchi S.
    (1981). The calmodulin binding protein in microtubules is tau factor. FEBS Lett 132, 137–140
    OpenUrlCrossRefPubMed
    1. Towbin H.,
    2. Staehelin T. and
    3. Gordon J.
    (1979). Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Nat. Acad. Sci. USA 76, 4350–4354
    OpenUrlAbstract/FREE Full Text
    1. Vale R. D.
    (1990). Microtubule-based motors proteins. Curr. Opin. Cell Biol 2, 15–22
    OpenUrlPubMed
    1. Vallee R. B. and
    2. Collins C. A.
    (1986). Purification of microtubules and microtubule associated proteins from sea urchins eggs and cultured mammalian cells using taxol, and use of exogenous stabilized brain microtubules for purifying microtubule associated proteins. Meth. Enzymol 134, 104–127
    OpenUrlCrossRefPubMedWeb of Science
    1. Venable J. and
    2. Coggeshall R.
    (1965). A simplified lead citrate stain for use in electron microscopy. J. Cell Biol 25, 407–.
    OpenUrlFREE Full Text
    1. Weingarten N. D.,
    2. Lokcwood A. H.,
    3. Hwo S. Y. and
    4. Kirschner N. W.
    (1975). A protein factor essential for microtubule assembly. Proc. Nat. Acad. Sci. USA 72, 1858–1862
    OpenUrlAbstract/FREE Full Text
    1. Wehland J.,
    2. Willingham M. C. and
    3. Sandoval I. V.
    (1983). A rat monoclonal antibody reacting specifically with the tyrosylated form of-tubulin. I. Biochemical characterization, effects on microtubule polymerization in vitro, and microtubule polymerization and organization in vivo. J. Cell Biol 97, 1467–1475
    OpenUrlAbstract/FREE Full Text
    1. Wolff A.,
    2. de Nechaud B.,
    3. Chillet D.,
    4. Mazarguil H.,
    5. Desbruyeres E.,
    6. Audebert S.,
    7. Edde B.,
    8. Gros F. and
    9. Denoulet P.
    (1992). Distribution of glutamylated-and -tubulin in mouse tissues using a specific monoclonal antibody, GT335. Eur. J. Cell Biol 59, 425–432
    OpenUrlPubMedWeb of Science
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Journal Articles
Stabilization and bundling of subtilisin-treated microtubules induced by microtubule associated proteins
Y. Saoudi, I. Paintrand, L. Multigner, D. Job
Journal of Cell Science 1995 108: 357-367;
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