Fig. 4

Apical IKNM Does Not Depend on Centrosome Integrity; CDK1 Activity Is Necessary and Sufficient for Apical IKNM

(A) Time-lapse imaging of a cell following laser ablation of the apical centrosome. (Left-most panel) Cell prior to ablation. The nucleus is labeled with PCNA-GFP (green). The membrane is visualized with Ras-GFP (green) and the centrosome with centrin-mKate2 (magenta, white arrow). After ablation (00:00), no centrin signal can be detected in the apical process (white arrow). The yellow box in panel 00:00 shows a 2× magnified region of the apical endfoot. The following panels show nuclear behavior following ablation marked by a yellow arrow. Time is in hr:min. The frames are from Movie S4. See also Figure S2.

(B–D) Time-lapse of cells expressing PCNA-GFP (gray) upon treatment with (B) DMSO only, (C) CDK1 inhibitor (RO3306), (D) Wee1 inhibitor (PD166285). Red arrows mark cells in S-phase (PCNA dots), and green arrows mark cells in G2 (disappearance of PCNA dots). Time is in hr:min. The frames are from Movie S5.

(E) Tracks of nuclear movements in the different drug conditions starting at the onset of apical IKNM (PD0166285), the entry into G2 phase (RO3306), or both (control).

(F) Average speed of apical nuclear migration in different conditions. Speed was calculated as total distance traveled by nuclei over time (µm/min). Error bars represent SEM.

Scale bars represent 10 µm. Apical surface is represented by a dotted line in (A) and a solid line in (B)–(D).