Cdc42 and mDia3 regulate microtubule attachment to kinetochores

Abstract

During mitosis, the mitotic spindle, a bipolar structure composed of microtubules (MTs) and associated motor proteins1,2, segregates sister chromatids to daughter cells. Initially some MTs emanating from one centrosome attach to the kinetochore at the centromere of one of the duplicated chromosomes. This attachment allows rapid poleward movement of the bound chromosome. Subsequent attachment of the sister kinetochore to MTs growing from the other centrosome results in the bi-orientation of the chromosome, in which interactions between kinetochores and the plus ends of MTs are formed and stabilized2. These processes ensure alignment of chromosomes during metaphase and their correct segregation during anaphase. Although many proteins constituting the kinetochore have been identified and extensively studied, the signalling responsible for MT capture and stabilization is unclear1,2. Small GTPases of the Rho family regulate cell morphogenesis by organizing the actin cytoskeleton and regulating MT alignment and stabilization3. We now show that one member of this family, Cdc42, and its effector, mDia3, regulate MT attachment to kinetochores.

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Figure 1: Effects of toxin B on mitosis.
Figure 2: Effects of Rho GTPase mutants on mitosis.
Figure 3: Interaction of mDia3 and CENP-A.
Figure 4: Effects of depletion of mDia3 on mitosis.

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Acknowledgements

We thank K. Aktories for Clostridium difficile toxin B, Y. Kiyosue for pQBI25-Xβ-tubulin, N. Mimori for CREST serum, S. Tsukita for the use of Delta-Vision system, T. Kiyomitsu, T. Tsuji, Y. Arakawa, J. Monypenny and N. Watanabe for advice, and M. Yanagida for discussion. This work was supported in part by Grants-in-aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan and from the Ministry of Health, Labour and Welfare of Japan.

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Correspondence to Shuh Narumiya.

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The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Methods and Figures (DOC 1467 kb)

41586_2004_BFnature02452_MOESM2_ESM.mov

Mitosis of control cells, merged images for GFP-Xbeta-tubulin and Hoechst 33342(Red). (MOV 801 kb)

Supplementary Movie 1

Mitosis of control cells, merged images for GFP-Xbeta-tubulin and Hoechst 33342(Red). (MOV 801 kb)

41586_2004_BFnature02452_MOESM3_ESM.mov

Mitosis of toxin B-treated cells, merged images for GFP-Xbeta-tubulin and Hoechst 33342(Red). (MOV 1063 kb)

Supplementary Movie 2

Mitosis of toxin B-treated cells, merged images for GFP-Xbeta-tubulin and Hoechst 33342(Red). (MOV 1063 kb)

41586_2004_BFnature02452_MOESM4_ESM.mov

Mitosis of control cells, merged images for GFP-EB1 and dsRed2-histoneH2B-k. (MOV 3862 kb)

Supplementary Movie 3

Mitosis of control cells, merged images for GFP-EB1 and dsRed2-histoneH2B-k. (MOV 3862 kb)

41586_2004_BFnature02452_MOESM5_ESM.mov

Mitosis of cells treated with mDia3 siRNA, merged images for GFP-EB1 and dsRed2-histoneH2B-k. (MOV 3072 kb)

Supplementary Movie 4

Mitosis of cells treated with mDia3 siRNA, merged images for GFP-EB1 and dsRed2-histoneH2B-k. (MOV 3072 kb)

41586_2004_BFnature02452_MOESM6_ESM.mov

Mitosis of cells treated with mDia3 siRNA, GFP-EB1 images. (MOV 3072 kb)

Supplementary Movie 5

Mitosis of cells treated with mDia3 siRNA, GFP-EB1 images. (MOV 3072 kb)

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Yasuda, S., Oceguera-Yanez, F., Kato, T. et al. Cdc42 and mDia3 regulate microtubule attachment to kinetochores. Nature 428, 767–771 (2004). https://doi.org/10.1038/nature02452

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