CC1–303-PH is defective for spindle oscillations but is otherwise sufficient for dynein-dependent spindle positioning. (A) The percentage of binucleated cells in cultures grown at 12°C for 16 h (n ≥ 495 cells for each strain). (B) The percentage of cells exhibiting stationary cortical Dyn1-3GFP foci (n ≥ 539 cells for each strain). (C) Cells expressing Dyn1-3GFP and mCherry-Tub1. Arrows indicate stationary cortical Dyn1-3GFP foci, and arrowheads indicate motile plus-end Dyn1-3GFP foci, as determined by two-color videos. Bars, 1 µm. (D) Box plot of cortical Dyn1-3GFP intensity. Foci in ΔTR and CC1–303-PH strains were not significantly different from those in wild-type NUM1 (P > 0.02 by a t test; n ≥ 20 foci for each strain). A.U., arbitrary unit. (E) Histograms of spindle penetration distance in HU-arrested NUM1 or CC1–303-PH cells in the kar9Δ background. Distance of spindle penetration, denoted by a red D, is defined as the farthest distance traveled by a spindle pole moving across the bud neck during a 10-min video (P = 0.0002 for CC1–303-PH vs. NUM1 by a t test; n ≥ 32 spindles for each strain from a single experiment). denotes mean ± SD. (F, left) The percentage of spindles in HU-arrested NUM1 kar9Δ or CC1–303-PH kar9Δ cells that crossed the bud neck over the course of a 10-min video (P < 0.0001 by a t test; n ≥ 181 for each strain from a single experiment). (right) Examples of GFP-Tub1 time-lapse images used for quantification. Time is in seconds. Arrowheads mark the bud neck. Bars, 1 µm. All error bars are standard error of proportion.
