Skip to main content
SpringerLink
Log in

The kinesin-immunoreactive homologue from Nicotiana tabacum pollen tubes: Biochemical properties and subcellular localization

  • Published:
Planta Aims and scope Submit manuscript

Abstract

In plant cells, microtubule-based motor proteins have not been characterized to the same degree as in animal cells; therefore, it is not yet clear whether the movement of organelles and vesicles is also dependent on the microtubular cytoskeleton. In this work the kinesinimmunoreactive homologue from pollen tubes of Nicotiana tabacum L. has been purified and biochemically characterized. The protein preparation mainly contained a polypeptide with a relative molecular weight of approx. 100 kDa. This polypeptide bound to animal microtubules in an ATP-dependent manner and it further copurified with an ATPase activity fourfold-stimulated by the presence of microtubules. In addition, the sedimentation coefficient (approx. 9S) was similar to those previously shown for other kinesins. Immunofluorescence analyses revealed a partial co-distribution of the protein with microtubules in the pollen tube. These data clearly indicate that several properties of the kinesin-immunoreactive homologue are similar to those of kinesin proteins, and suggest that molecular mechanisms analogous to those of animal cells may drive the microtubule-based motility of organelles and vesicles in plants.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

AE-LPLC:

anion-exchange low-pressure liquid chromatography

AMPPNP:

5′-adenylylimidodiphosphate

PKH:

pollen kinesin homologue

SDS-PAGE:

sodium dodecyl sulfate polyacrylamide gel electrophoresis

References

  • Amos, L.A. (1987) Kinesin from pig brain studied by electron microscopy. J. Cell Sci. 87, 105–111

    Google Scholar 

  • Asada, T., Sonobe, S., Shibaoka, H. (1991) Microtubule translocation in the cytokinetic apparatus of cultured tobacco cells. Nature 350, 238–241

    Google Scholar 

  • Baxter-Gabbard, K.L. (1972) A simple method for the large-scale preparation of sucrose gradients. FEBS Lett. 20, 117–119

    Google Scholar 

  • Bloom, G.S. (1992) Motor proteins for cytoplasmic microtubules. Curr. Opin. Cell Biol. 4, 66–73

    Google Scholar 

  • Bloom, G.S., Wagner, M.C., Pfister, K.K., Brady, S.T. (1988) Native structure and physical properties of bovine brain kinesin and identification of the ATP-binding subunit polypeptide. Biochemistry 27, 3409–3416

    Google Scholar 

  • Bradford, M.M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein using the principle of protein-dye binding. Anal. Biochem. 72, 248–254

    Article  CAS  PubMed  Google Scholar 

  • Brady, S.T., Pfister, K.K., Bloom, G.S. (1990) A monoclonal antibody to the heavy chain of kinesin inhibits anterograde and retrograde axonal transport in isolated squid axoplasm. Proc. Natl. Acad. Sci. USA 87, 1061–1065

    Google Scholar 

  • Brewbaker, J.L., Kwack, B.H. (1963) The essential role of calcium ions in pollen germination and pollen tube growth. Am. J. Bot. 50, 859–865

    Google Scholar 

  • Buster, D., Scholey, J.M. (1991) Purification and assay of kinesin from sea urchin eggs and early embryos. J. Cell Sci. 14, 109–115

    Google Scholar 

  • Cohn, S.A., Ingold, A.I., Scholey, J.M. (1987) Correlation between the ATPase and microtubule translocating activities of sea urchin egg kinesin. Nature 328, 160–163

    Google Scholar 

  • Cole, D.G., Cande, W.Z., Baskin, R.J., Skoufias, D.A., Hogan, C.J., Scholey, J.M. (1992) Isolation of sea urchin egg kinesin-related protein using peptide antibodies. J. Cell Sci. 101, 291–301

    Google Scholar 

  • Dabora, S.L., Sheetz, M.P. (1988) The microtubule-dependent formation of a tubulovesicular network with characteristics of the ER from cultured cell extracts. Cell 54, 27–35

    Google Scholar 

  • De Cuevas, M., Tao, T, Goldstein, L.S.B. (1992) Evidence that the stalk of Drosophila kinesin heavy chain is an α-helical coiled coil. J. Cell Biol. 116, 957–965

    Google Scholar 

  • Emons, A.M.C., Pierson, E.S., Derksen, J. (1991) Cytoskeleton and intracellular movement in plant cells. In: Biotechnology — current progress, pp. 311–335, Cheremisimoff, P.N., Ferrante, L.M., eds. Technomic Publishing Co, Lancaster, Basel

    Google Scholar 

  • Epstein, H., Scholey, J.M. (1992) Kinesins in the spindle: an update. Trends Cell Biol. 2, 315–318

    Google Scholar 

  • Fellous, A., Francon, J., Lennon, A.M., Nunez, J. (1977) Microtubule assembly in vitro. Eur. J. Biochem. 78, 167–174

    Google Scholar 

  • Fosket, D.E., Morejohn, L.C. (1992) Structural and functional organization of tubulin. In: Annu. Rev. Plant Physiol. Plant Mol. Biol. 43, 201–240

    Google Scholar 

  • Gonzalez-Romo, P., Sanchez-Nito, S., Gavilanes-Ruiz, M. (1992) A modified colorimetric method for the determination of orthophosphate in the presence of high ATP concentrations. Anal. Biochem. 200, 235–238

    Google Scholar 

  • Hepler, P.K., Palevitz, B.A., Lancelle, S.A., McCauley, M.M., Lichtscheidl, I. (1990) Cortical endoplasmic reticulum in plants. J. Cell Sci. 96, 355–373

    Google Scholar 

  • Hirokawa, N.N., Pfister, K.K., Yorufuji, H., Wagner, M.C., Brady, S.T., Bloom, G.S. (1989) Submolecular domains of bovine brain kinesin identified by electron microscopy and monoclonal antibody decoration. Cell 56, 867–878

    Google Scholar 

  • Hirokawa, N., Sato-Yoshitake, R., Kobayashi, N., Pfister, K.K., Bloom, G.S., Brady, S.T. (1991) Kinesin associates with anterogradely transported membranous organelles in vivo. J. Cell Biol. 114, 295–302

    Google Scholar 

  • Hollenbeck, P.J. (1989) The distribution, abundance and subcellular localization of kinesin. J. Cell Biol. 108, 2335–2342

    Google Scholar 

  • Hollenbeck, P.J., Swanson, J.A. (1990) Radial extension of the macrophage tubular lysosomes supported by kinesin. Nature 346, 864–866

    Google Scholar 

  • Houliston, E., Elinson, R.P. (1991) Evidence for the involvement of microtubules, ER, and kinesin in the cortical rotation of fertilized frog eggs. J. Cell Biol. 114, 1017–1028

    Google Scholar 

  • Ingold, A.L., Cohn, S.A., Scholey, J.M. (1988) Inhibition of kinesindriven microtubule motility by monoclonal antibodies to kinesin heavy chains. J. Cell Biol. 107, 2657–2667

    Google Scholar 

  • Kachar, B., Albanesi, J.P., Fujisaki, M., Korn, E.D. (1987) Extensive purification from Acanthamoeba castellanii of a microtubule-dependent translocator with microtubule-activated Mg2+-ATPase activity. J. Biol. Chem. 262, 16180–16185

    Google Scholar 

  • Khono, T., Ishikawa, R., Negata, T., Kohama, K., Shimmen, T. (1992) Partial purification of myosin from lily pollen tubes by monitoring with in vitro motility assay. Protoplasma 170, 77–85

    Google Scholar 

  • Kuznetsov, S.A., Vaisberg, Y.A., Shanina, N.A., Magretova, N.N., Chernyak, V.Y., Gelfand, V.I. (1988) The quaternary structure of bovine brain kinesin. EMBO J. 7, 353–356

    Google Scholar 

  • Kuznetsov, S.A., Vaisberg, Y.A., Rothwell, S.W., Murphy, D.B., Gelfand, V.I. (1989) Isolation of a 45-kDa fragment from the kinesin heavy chain with enhanced ATPase and microtubulebinding activities. J. Biol. Chem. 264, 589–595

    Google Scholar 

  • Laemmli, U.K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685

    PubMed  Google Scholar 

  • Leslie, R.J., Hird, R.B., Wilson, L., McIntosh, J.R., Scholey, J.M. (1987) Kinesin is associated with a nonmicrotubule component of sea urchin mitotic spindle. Proc. Natl. Acad. Sci. USA 84, 2771–2774

    Google Scholar 

  • Leopold, P.L., McDowall, A.W., Pfister, K.K., Bloom, G.S., Brady, S.T. (1992) Association of kinesin with characterized membrane-bounded organelles. Cell Motil. Cytoskel. 23, 19–33

    Google Scholar 

  • Martin, R.G., Ames, B.N. (1961) A method for determining the sedimentation behavior of enzymes: application to protein mixtures. J. Biol. Chem. 236, 1372–1379

    Google Scholar 

  • McCaffrey, G., Vale, R.D. (1989) Identification of a kinesin-like microtubule-based motor protein in Dyctiostelium discoideum. EMBO J. 8, 3229–3234

    Google Scholar 

  • Murofushi, H., Ikai, A., Okuhara, K., Kotani, S., Aizawa, H., Kumakura, K., Sakai, H. (1988) Purification and characterization of kinesin from adrenal medulla. J. Biol. Chem. 263, 12744–12750

    Google Scholar 

  • Neighbors, B.W., Williams, R.C.Jr., McIntosh, J.R. (1988) Localization of kinesin in cultured cells. J. Cell Biol. 106, 1193–1204

    Google Scholar 

  • Pfister, K.K., Wagner, M.C., Stenoien, D.L., Brady, S.T., Bloom, G.S. (1989) Monoclonal antibodies to kinesin heavy chains stain vesicle-like structures, but not microtubules in cultured cells. J. Cell Biol. 108, 1453–1463

    Google Scholar 

  • Pierson, E.S., Cresti, M. (1992) Cytoskeleton and cytoplasmic organization of pollen and pollen tube. Int. Rev. Cytol. 140, 73–125

    Google Scholar 

  • Rodionov, V.I., Gyoeva, F.K., Gelfand, V.I. (1991) Kinesin is responsible for centrifugal movement of pigment granules in melanophores. Proc. Natl. Acad. Sci. USA 88, 4956–4960

    Google Scholar 

  • Saxton, W.M., Porter, M.A., Cohn, S.A., Scholey, J.M., Raff, E.M., McIntosh, J.R. (1988) Drosophila kinesin: characterization of microtubule motility and ATPase. Proc. Natl. Acad. Sci. USA 85, 1109–1113

    Google Scholar 

  • Saxton, W.M., Hicks, J., Goldstein, L.S.B., Raff, E.C. (1991) Kinesin heavy chain is essential for viability and neuromuscular functions in Drosophila, but mutants show no defects in mitosis. Cell 64, 1093–1102

    Google Scholar 

  • Scholey, J.M., Porter, M.E., Grissom, P.M., McIntosh, J.R. (1985) Identification of kinesin in sea urchin eggs and evidence for its localization in the mitotic spindle. Nature 318, 483–486

    Google Scholar 

  • Scholey, J.M., Heuser, J., Yang, J.T., Goldstein, L.S.B. (1989) Identification of the globular mechanochemical heads of kinesin. Nature 338, 355–357

    Google Scholar 

  • Schroer, T.A., Schnapp, B.J., Reese, T.S., Sheetz, M.P. (1988) The role of kinesin and other soluble factors in organelle movement along microtubules. J. Cell Biol. 107, 1785–1792

    Google Scholar 

  • Tiezzi, A. (1991) The pollen tube Cytoskeleton. Electron Microsc. Rev. 4, 205–219

    Google Scholar 

  • Tiezzi, A., Moscatelli, A., Milanesi, C., Ciampolini, F., Cresti, M. (1987) Taxol-induced structures derived from cytoskeletal elements of Nicotiana tabacum pollen tube. J. Cell Sci. 88, 657–661

    Google Scholar 

  • Tiezzi, A., Moscatelli, A., Cai, G., Bartalesi, A., Cresti, M. (1992) An immunoreactive homologue of mammalian kinesin in Nicotiana tabacum pollen tube. Cell Motil. Cytoskel. 21, 132–137

    Google Scholar 

  • Towbin, H., Staehelin, T., Gordon, J. (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets. Procedure and some applications. Proc. Natl Acad. Sci. USA 76, 4350–4354

    Google Scholar 

  • Vale, R.D., Reese, T.S., Sheetz, M.P. (1985) Identification of a novel force-generating protein, kinesin, involved in microtubule-based motility. Cell 42, 39–50

    Google Scholar 

  • Vallee, R.B., Shpetner, H.S. (1990) Motor proteins of cytoplasmic microtubules. Annu. Rev. Biochem. 59, 909–932

    Google Scholar 

  • Wright, B.D., Wedaman, K.P., Willy, P.J., Morand, J.N., Scholey, J.M. (1991) Subcellular localization and sequence of sea urchin kinesin heavy chain: evidence for its association with membranes in the mitotic apparatus and interphase cytoplasm. J. Cell Biol. 113, 817–833

    Google Scholar 

  • Wagner, M.C., Pfister, K.K., Bloom, G.S., Brady, S.T. (1989) Copurification of kinesin polypeptides with microtubule-stimulated Mg-ATPase activity and kinetic analysis of enzymatic properties. Cell Motil. Cytoskel. 12, 195–215

    Google Scholar 

  • Yang, J.T., Saxton, W.M., Stewart, R.J., Raff, E.C., Golstein, L.S.B. (1990) Evidence that the head of kinesin is sufficient for force generation and motility in vitro. Science 249, 42–47

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Dipartimento Biologia Ambientale, Università di Siena, Via Mattioli 4, I-53100, Siena, Italy

    G. Cai, A. Bartalesi, C. Del Casino, A. Moscatelli, A. Tiezzi & M. Cresti

Authors
  1. G. Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A. Bartalesi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. C. Del Casino
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. A. Moscatelli
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. A. Tiezzi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M. Cresti
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cai, G., Bartalesi, A., Del Casino, C. et al. The kinesin-immunoreactive homologue from Nicotiana tabacum pollen tubes: Biochemical properties and subcellular localization. Planta 191, 496–506 (1993). https://doi.org/10.1007/BF00195751

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00195751

Key words

Search

Navigation