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Published 24 October 2005. doi:10.1083/jcb.200505107
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 171, Number 2, 229-240
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Article

 Mechanisms for focusing mitotic spindle poles by minus end–directed motor proteins

Gohta Goshima1, François Nédélec2, and Ronald D. Vale1

1 The Howard Hughes Medical Institute and the Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94107
2 The European Molecular Biology Laboratory, Cell Biology and Biophysics Programme, 69117 Heidelberg, Germany

Correspondence to Ron Vale: vale{at}cmp.ucsf.edu; or François Nédélec: nedelec{at}embl.de

During the formation of the metaphase spindle in animal somatic cells, kinetochore microtubule bundles (K fibers) are often disconnected from centrosomes, because they are released from centrosomes or directly generated from chromosomes. To create the tightly focused, diamond-shaped appearance of the bipolar spindle, K fibers need to be interconnected with centrosomal microtubules (C-MTs) by minus end–directed motor proteins. Here, we have characterized the roles of two minus end–directed motors, dynein and Ncd, in such processes in Drosophila S2 cells using RNA interference and high resolution microscopy. Even though these two motors have overlapping functions, we show that Ncd is primarily responsible for focusing K fibers, whereas dynein has a dominant function in transporting K fibers to the centrosomes. We also report a novel localization of Ncd to the growing tips of C-MTs, which we show is mediated by the plus end–tracking protein, EB1. Computer modeling of the K fiber focusing process suggests that the plus end localization of Ncd could facilitate the capture and transport of K fibers along C-MTs. From these results and simulations, we propose a model on how two minus end–directed motors cooperate to ensure spindle pole coalescence during mitosis.

Abbreviations used in this paper: BA, bleached area; C-MT, centrosomal microtubule; NBA, nonbleached area; NES, nuclear export signals; RNAi, RNA interference.


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