Relocalization of Kinesin-6 to microtubule tips and the equatorial cortex during anaphase. S2 cells coexpressing mCherry-tubulin (red) and Kinesin-6–GFP (green) were imaged by dual-color TIRF microscopy. (A) At the 0 min time point, the cell is in metaphase, and subsequent times are in anaphase. The inset at 6 min shows Kinesin-6–GFP colocalizing with the tips of microtubules. A diffuse equatorial band of Kinesin-6–GFP is also seen at this time point. At later time points, Kinesin-6–GFP is localized exclusively to the equator, which likely contains antiparallel microtubule bundles. Although the total Kinesin-6–GFP signal appears to diminish over time, the majority of the protein concentrates on the microtubules in the interior central spindle (Goshima and Vale, 2005), which is not visible or minimally visible by TIRF microscopy. See Video 5. Bar, 10 µm. (B) This time series from another cell shows microtubules that grow toward one another and increase levels of Kinesin-6–GFP when the microtubules come in close proximity (arrowheads), including in regions where the microtubules are not directly overlapping (40 s). Bar, 2 µm.