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Protoplasma

, Volume 173, Issue 1–2, pp 77–85 | Cite as

Microtubule-nucleation sites on nuclei of higher plant cells

  • K. Mizuno
Article

Summary

The nucleation and the elongation of microtubules from isolated nuclei of higher plant cells were investigated. Isolated intact nuclei failed to nucleate microtubules at their surface when they were incubated with purified tubulin from plant or animal sources. However, frozen and thawed nuclei or nuclear particles obtained by gentle nuclei homogenization nucleated microtubules and nucleated microtubules elongated radially from the surface of nuclei or from the nuclear particles. Microtubules radiating from the nuclear particles were very much shorter than those radiating from frozen and thawed nuclei. The washing of the nuclear particles diminished the ability of the particles to nucleate microtubules. The ability of the washed nuclear particles to nucleate microtubules was restored by the addition of the soluble fraction of a nuclear homogenate. The complexes of radiating microtubules could easily be observed under a phasecontrast microscope. Electron microscopy demonstrated that microtubules in the complexes formed bundles. The staining with a monoclonal antibody specific for plant tubulin of the complexes of radiating microtubules, prepared by successive polymerization of animal tubulin and plant tubulin, revealed that microtubules in the complex incorporated tubulin at their proximal ends. This result indicates that the mode of incorporation of tubulin onto frozen and thawed nuclei or onto the nuclear particles is different from that in pericentriolar bodies in animal cells. Mg2+ seems to participate in the regulatory mechanism that determines the length of microtubules on the complexes.

Keywords

Higher plant nuclei Microtubule-nucleation sites Microtubule-organizing center Tubulin-incorporating ends 

Abbreviations

MTOC

microtubule-organizing center

MES

2-(N-morpholino) ethane-sulfonic acid

PIPES

piperazine-N,N′-bis(2-ethanesulfonic acid)

PMSF

phenylmethyl sulfonyl fluoride

EDTA

ethylenediaminetetraacetic acid

EGTA

ethylene glycol bis (β-aminoethylether)-N,N,N′,N′-tetraacetic acid

GTP

guanosine triphosphate

NP-40

Nonidet P-40

DMSO

dimethylsulfoxide

EPC

ethyl N-phenylcarbamate

SDS-PAGE

sodium dodecyl sulfate-polyacrylamide gel electrophoresis

DAPI

4′,6-diamidiho-2-phenylindole

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Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • K. Mizuno
    • 1
  1. 1.Department of Biology, Faculty of ScienceOsaka UniversityOsakaJapan

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