Dynein is necessary for ternary complex–induced spindle oscillations in HeLa cells. (A–F) Images from time-lapse microscopy of metaphase HeLa Kyoto cells stably expressing GFP–α-tubulin as well as mCherry-H2B and transfected with YFP (A), Gαi₁-YFP (B), YFP-LGN (C), Gαi₁-YFP and p150^Glued siRNAs (D), YFP-LGN and p150^Glued siRNAs (E), or Gαi₁-Δmyr-YFP (F; see also Videos 1–4). Note that cortical YFP-LGN, which is present at low levels, is not visible in C and E because of the strong GFP–α-tubulin signal but can be seen better in Video 1 (S4), in which YFP-LGN is transfected in HeLa Kyoto cells expressing solely mCherry-H2B. In this and other figures with time-lapse sequences, the position of chromosomes is indicated by a white line when chromosomes are within the imaging plane and by an asterisk when the metaphase plate is no longer in the plane of view. 10 cells were imaged for each condition. The bar graphs on the right are readouts of the extent of spindle oscillations, representing the frequency at which chromosome position changes >10° between two frames (including when they move out of the imaging plane), along with the SEM. Two-tailed Student’s t tests show that the extent of spindle oscillations upon overexpression of Gαi₁-YFP or of YFP-LGN is statistically different from that observed in control conditions; in both cases, P < 0.0001. Similarly, the values in Gαi₁-YFP and p150^Glued siRNA (D), as well as in YFP-LGN and p150^Glued siRNA (E), are statistically different from those of Gαi₁-YFP (B) or YFP-LGN (C) overexpression alone; in both cases, P < 0.0005. In contrast, the values in Gαi₁-YFP and p150^Glued siRNA (D), as well as in YFP-LGN and p150^Glued siRNA (E), are not statistically different from those in the control condition (A); P = 0.16 and P = 0.18, respectively. Time is indicated in hours and minutes.