Aster centration is driven by MT-pulling forces in the cytoplasm. (A) Time-dependent immunostaining of centering MT asters (MT, green; DNA, red). The indicated time is taken in reference with sperm entry by accounting for a mean 3-min delay between sperm addition and entry. (B) Centering MT asters are ablated parallel to the centration trajectory. A drift in the trajectory toward (away from) the ablation line suggests that MTs are pushing (pulling). (C) In situ MT aster immunostaining performed immediately after laser ablation along the red broken line. (D and E) Time-lapse (D) and time projection (E) of a centering aster ablated as indicated. Time 0 is the time of ablation. White arrowhead, male pronucleus. (F) Definition of aster velocity and drift after ablation. (G) Aster drift in the indicated conditions (n = 10 for control; n = 28 for side ablation; n = 7 for nocodazole; n = 10 for nocodazole + side ablation). Bar, 10 μm. (H) Aster velocity vectors after ablation at the indicated locations. (I) Aster speed along the centration path after ablation in the indicated conditions. (J) Distance–time plot of a front-ablated aster. (K) Aster velocity along the centration path after ablation (V2) as a function of the velocity before (V1). n.s., nonsignificant; **, P < 10−4 (Student’s t test). Error bars represent SD. Bars, 50 µm.
