Dynein dephosphorylation requires MT attachment and tension. (A) IFM analysis of methanol-fixed NRK2 cells stained for chromosomes (DAPI; 1, 5, 9, and 13), phosphodynein (PT89; 2, 6, 10, and 14), and BubR1 (3, 7, 11, and 15) was performed after the following treatments: untreated (1–4), nocodazole (5–8), taxol (9–12), and calyculin A (13–16). Methanol fixation preserves kinetochore- but not spindle-associated BubR1. (B) NRK2 cells were transfected with GFP-tagged wild-type (1–4) or YFP-tagged H125A mutant PP1-γ (5–8) and stained for chromosomes (1 and 5), PT89 (2 and 6), or BubR1 (3 and 7). (A and B) Scales display fluorescence intensity ranges of 1–1,000 intensity units for PT89 and 0–1,200 intensity units for BubR1. Bars, 5 μm.