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Vol. 13, Issue 12, 4308-4316, December 2002
§ and
*Department of Cell Biology, Harvard Medical School, Boston,
Massachusetts 02115; Microtubule polymerization dynamics at kinetochores is
coupled to chromosome movements, but its regulation there is poorly understood. The plus end tracking protein EB1 is required both for
regulating microtubule dynamics and for maintaining a euploid genome.
To address the role of EB1 in aneuploidy, we visualized its targeting
in mitotic PtK1 cells. Fluorescent EB1, which localized to polymerizing
ends of astral and spindle microtubules, was used to track their
polymerization. EB1 also associated with a subset of attached
kinetochores in late prometaphase and metaphase, and rarely
in anaphase. Localization occurred in a narrow crescent, concave toward
the centromere, consistent with targeting to the microtubule plus
end-kinetochore interface. EB1 did not localize to
kinetochores lacking attached kinetochore
microtubules in prophase or early prometaphase, or upon nocodazole
treatment. By time lapse, EB1 specifically targeted to
kinetochores moving antipoleward, coupled to microtubule
plus end polymerization, and not during plus end depolymerization. It
localized independently of spindle bipolarity, the spindle checkpoint,
and dynein/dynactin function. EB1 is the first protein whose targeting
reflects kinetochore directionality, unlike other plus end
tracking proteins that show enhanced kinetochore binding in
the absence of microtubules. Our results suggest EB1 may modulate
kinetochore microtubule polymerization and/or attachment.
Department of Biology, University
of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599;
and §Marine Biological Laboratory, Woods Hole,
Massachusetts 02543
Online
version of this article contains video material for some figures.
Online version available at www.molbiolcell.org.
Corresponding author. E-mail address:
jennifer_tirnauer{at}hms.harvard.edu.
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