Journal Article

Isolation of a Novel 190 kDa Protein from Tobacco BY-2 Cells: Possible Involvement in the Interaction between Actin Filaments and Microtubules

Hisako Igarashi, Hidefumi Orii, Hitoshi Mori, Teruo Shimmen and Seiji Sonobe

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 41, issue 8, pages 920-931
Published in print July 2000 | ISSN: 0032-0781
Published online July 2000 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcd015
Isolation of a Novel 190 kDa Protein from Tobacco BY-2 Cells: Possible Involvement in the Interaction between Actin Filaments and Microtubules

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  • Biochemistry
  • Molecular and Cell Biology
  • Plant Sciences and Forestry

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Interaction between actin filaments (AFs) and microtubules (MTs) has been reported in various plant cells, and the presence of a factor(s) connecting these two cytoskeletal networks has been suggested, but its molecular entity has not been elucidated yet. We obtained a fraction containing MT-binding polypeptides, which induced bundling of AFs and of MTs. A 190 kDa polypeptide which associated with AFs was selectively isolated from the fraction. This polypeptide was thought to have an ability to bind to both AFs and MTs. We raised a monoclonal antibody against the 190 kDa polypeptide. Immunostaining demonstrated the association of the 190 kDa polypeptide with AF bundles and with MT bundles formed in vitro. Immunocytochemical studies throughout the cell cycle revealed that the 190 kDa polypeptide was localized in the nucleus before nuclear envelope breakdown, and in the spindle and the phragmoplast during cell division. After the re-formation of the nuclear envelope, the 190 kDa polypeptide was sequestered to the daughter nuclei. Using the antibody, we succeeded in cloning a cDNA encoding the 190 kDa polypeptide.

Keywords: Key words: Actin filaments — Microtubules — Microtubule-binding proteins.; Abbreviations: AF(s), actin filament(s); MT(s), microtubule(s); DMSO, dimethyl sulfoxide; DTT, dithiothreitol; PMSF, phenylmethanesulfuoride; EGTA, ethyleneglycol-bis-(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid; PIPES, piperazine-N,N′-bis-(2-ethanaesulfonic acid); PBS, phosphate buffered saline; TCA, trichloroacetic acid.

Journal Article.  5877 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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