LOCALIZATION OF THE KINESIN-LIKE PROTEIN XKLP2 TO SPINDLE POLES REQUIRES A LEUCINE-ZIPPER, A MICROTUBULE-ASSOCIATED PROTEIN, AND DYNEIN

Xklp2 is a plus end-directed Xenopus kinesin-like protein localized at spindle poles and required for centrosome separation during spindle a ssembly in Xenopus egg extracts. A glutathione-S-transferase fusion pr otein containing the COOH-terminal domain of Xklp2 (GST-Xklp2-Tail) wa s previously found to localize to spindle poles (Boleti, H., E. Karsen ti, and I. Vernos. 1996. Cell. 84:49-59). Now, we have examined the me chanism of localization of GST-Xklp2-Tail. Immunofluorescence and elec tron microscopy showed that Xklp2 and GST-Xklp2-Tail localize specific ally to the minus ends of spindle pole and aster microtubules in mitot ic, but not in interphase, Xenopus egg extracts. We found that dimeriz ation and a COOH-terminal leucine zipper are required for this localiz ation: a single point mutation in the leucine zipper prevented targeti ng. The mechanism of localization is complex and two additional factor s in mitotic egg extracts are required for the targeting of GST-Xklp2- Tail to microtubule minus ends: (a) a novel 100-kD microtubule-associa ted protein that we named TPX2 (Targeting protein for Xklp2) that medi ates the binding of GST-Xklp2-Tail to microtubules and (b) the dynein- dynactin complex that is required for the accumulation of GST-Xklp2-Ta il at microtubule minus ends. We propose two molecular mechanisms that could account for the localization of Xklp2 to microtubule minus ends .