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© The Rockefeller University Press, 0021-9525/1998//673 $5.00
The Journal of Cell Biology, Volume 143, Number 3, , 1998 673-685

Regular Articles

Localization of the Kinesin-like Protein Xklp2 to Spindle Poles Requires a Leucine Zipper, a Microtubule-associated Protein, and Dynein



Torsten Wittmann*, Haralabia Boleti{ddagger}, Claude Antony§, Eric Karsenti*, and Isabelle Vernos*

* European Molecular Biology Laboratory, Cell Biology and Cell Biophysics Programs, D-69117 Heidelberg, Germany; {ddagger} Institut Pasteur, 75724 Paris Cedex 15, France; and § Institut Curie, 75248 Paris Cedex 05, France

Xklp2 is a plus end–directed Xenopus kinesin-like protein localized at spindle poles and required for centrosome separation during spindle assembly in Xenopus egg extracts. A glutathione-S-transferase fusion protein containing the COOH-terminal domain of Xklp2 (GST-Xklp2-Tail) was previously found to localize to spindle poles (Boleti, H., E. Karsenti, and I. Vernos. 1996. Cell. 84:49–59). Now, we have examined the mechanism of localization of GST-Xklp2-Tail. Immunofluorescence and electron microscopy showed that Xklp2 and GST-Xklp2-Tail localize specifically to the minus ends of spindle pole and aster microtubules in mitotic, but not in interphase, Xenopus egg extracts. We found that dimerization and a COOH-terminal leucine zipper are required for this localization: a single point mutation in the leucine zipper prevented targeting. The mechanism of localization is complex and two additional factors in mitotic egg extracts are required for the targeting of GST-Xklp2-Tail to microtubule minus ends: (a) a novel 100-kD microtubule-associated protein that we named TPX2 (Targeting protein for Xklp2) that mediates the binding of GST-Xklp2-Tail to microtubules and (b) the dynein–dynactin complex that is required for the accumulation of GST-Xklp2-Tail at microtubule minus ends. We propose two molecular mechanisms that could account for the localization of Xklp2 to microtubule minus ends.

Key Words: kinesin-like protein • microtubule-associated protein • spindle • microtubule • dynein

Abbreviations used in this paper: GST, glutathione-S-transferase; KLP, kinesin-like protein; MAP, microtubule-associated protein; Tm, melting temperature; TPX2, Targeting protein for Xklp2.

Address all correspondence to Isabelle Vernos, EMBL Heidelberg, Meyerhofstrasse 1, D-69117 Heidelberg, Germany. Tel.: 49 6221 387 324. Fax: 49 6221 387 306. E-mail: vernos{at}embl-heidelberg.de


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