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Localization of actin filaments on mitotic apparatus in tobacco BY-2 cells

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Abstract

Actin filaments are among the major components of the cytoskeleton, and participate in various cellular dynamic processes. However, conflicting results had been obtained on the localization of actin filaments on the mitotic apparatus and their participation in the process of chromosome segregation. We demonstrated by using rhodamine-phalloidin staining, the localization of actin filaments on the mitotic spindles of tobacco BY-2 cells when the cells were treated with cytochalasin D. At prophase, several clear spots were observed at or near the kinetochores of the chromosomes. At anaphase, the actin filaments that appeared to be pulling chromosomes toward the division poles were demonstrated. However, as there was a slight possibility that these results might have been the artifacts of cytochalasin D treatment or the phalloidin staining, we analyzed the localization of actin filaments at the mitotic apparatus immunologically. We cloned a novel BY-2 α-type actin cDNA and prepared a BY-2 actin antibody. The fluorescence of the anti-BY-2 actin antibody was clearly observed at the mitotic apparatus in both non-treated and cytochalasin D-treated BY-2 cells during mitosis. The facts that similar results were obtained in both actin staining with rhodamine-phalloidin and immunostaining with actin antibody strongly indicate the participation of actin in the organization of the spindle body or in the process of chromosome segregation. Furthermore, both filamentous actin and spindle bodies disappeared in the cells treated with propyzamide, which depolymerizes microtubules, supporting the notion that actin filaments are associated with microtubules organizing the spindle body.

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Acknowledgments

The authors thank Dr. T Nakano of Osaka Prefecture University for technical support in the preparation of BY-2 actin antibody.

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Authors and Affiliations

  1. Cryobiosystem Research Center, Iwate University, Morioka, Iwate, 020-8550, Japan

    Hiroshi Yasuda, Katsuhiro Kanda, Kayoko Suenaga, Shin-ichiro Kidou & Shin-ichiro Ejiri

  2. Iwate Biotechnology Research Center, Kitakami, Iwate, 024-0003, Japan

    Hiroyuki Koiwa

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  1. Hiroshi Yasuda
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  2. Katsuhiro Kanda
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  3. Hiroyuki Koiwa
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Correspondence to Shin-ichiro Ejiri.

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Hiroshi Yasuda and Katsuhiro Kanda contributed equally.

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Yasuda, H., Kanda, K., Koiwa, H. et al. Localization of actin filaments on mitotic apparatus in tobacco BY-2 cells. Planta 222, 118–129 (2005). https://doi.org/10.1007/s00425-005-1522-8

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