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Kinetochore dynein: its dynamics and role in the transport of the Rough deal checkpoint protein

Nature Cell Biology volume 3, pages 1001–1007 (2001)Cite this article

Abstract

We describe the dynamics of kinetochore dynein–dynactin in living Drosophila embryos and examine the effect of mutant dynein on the metaphase checkpoint. A functional conjugate of dynamitin with green fluorescent protein accumulates rapidly at prometaphase kinetochores, and subsequently migrates off kinetochores towards the poles during late prometaphase and metaphase. This behaviour is seen for several metaphase checkpoint proteins, including Rough deal (Rod). In neuroblasts, hypomorphic dynein mutants accumulate in metaphase and block the normal redistribution of Rod from kinetochores to microtubules. By transporting checkpoint proteins away from correctly attached kinetochores, dynein might contribute to shutting off the metaphase checkpoint, allowing anaphase to ensue.

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Figure 1: p50/Dmn–GFP protein co-fractionates with the dynactin complex (19S) on a sucrose density gradient.
Figure 2: Dmn–GFP dynamics in living embryos.
Figure 3: Dynein and Rod colocalize during mitosis in Drosophila neuroblasts.
Figure 4: Rod localization at metaphase is dependent on dynein function.
Figure 5: Mutant dynein accumulates on kinetochores and does not spread polewards along kinetochore microtubules in metaphase third-instar giant neuroblasts.
Figure 6: Drugs that alter microtubule assembly and dynamics block polewards migration of dynein.
Figure 7: Dynein mutant neuroblasts are delayed in metaphase and suffer mitotic defects, but do not exhibit premature sister chromatid separation.
Figure 8: Dynein mutants exhibit defects in spindle assembly that might affect chromosome behaviour and checkpoint controls.

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Acknowledgements

We acknowledge helpful discussions with all members of the Hays laboratory, and thank E. D. Salmon for communicating results before publication. This work was supported by grants to T.H. and E.W. from the National Institutes of Health (GM44757 and GM19123 respectively) and the American Heart Association (96002200). R.K., R.B. and F.S. performed this work at the UPR 2420 Centre de Génétique Moléculaire of the CNRS, associated with the Université Pierre et Marie Curie. R.K. was supported in part by grants from the CNRS and from the Association Pour la Recherche sur le Cancer (France). R.B. was supported by grants BD/11488/97 and P/BIA/111055/1998 from FCT, Portugal. F.S. was supported by Le Ministère Nationale de l'Enseignement Supérieur et de la Recherche.

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Authors and Affiliations

  1. Department of Biology, Virginia Tech University, Blacksburg, 24061, Virginia, USA

    Edward Wojcik

  2. Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, 55108, Minnesota, USA

    Edward Wojcik, Madeline Serr & Thomas Hays

  3. C.N.R.S., Centre de Génétique Moleculaire, Avenue de la Terrasse, Gif-sur-Yvette, 91198, France

    Renata Basto, Frédéric Scaërou & Roger Karess

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Correspondence to Edward Wojcik.

Supplementary information

Figure S1

Dynein mutant neuroblasts retain a checkpoint that prevents pre-mature degradation of cyclin B. (PDF 76 kb)

Movie 1

Time-lapse recordings of transformed syncytial were obtained by confocal microscopy, with 5 second intervals between frames. (MOV 2135 kb)

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Wojcik, E., Basto, R., Serr, M. et al. Kinetochore dynein: its dynamics and role in the transport of the Rough deal checkpoint protein. Nat Cell Biol 3, 1001–1007 (2001). https://doi.org/10.1038/ncb1101-1001

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