Hook2 localizes to the central spindles and promotes recruitment of dynein and dynactin to the central spindles. (A and B) Representative images of primary MEFs (A) and HeLa cells (B) in indicated stages of the cell cycle stained for endogenous Hook2, MTs with α-tubulin, and chromosomes with DAPI. Bars, 10 µm. (C) Western blot showing enrichment of Hook2 in midbody pellet from synchronized HeLa cells as compared with supernatant containing cytosol from the same cells. (D–G) Representative images of HeLa cells in anaphase treated with indicated siRNA. MTs were stained with α-tubulin to visualize spindles. Dynein, dynactin, and spindle midzone were visualized by antibodies against DIC, p150^glued, CYK4 and MKLP1, respectively, and chromosomes were visualized by DAPI in each case. Bars, 10 µm. (H–L) Quantification of the relative levels (control siRNA) of α-tubulin, p150^glued, DIC, MKLP1, and CYK4 at central spindles and spindle midzone, respectively (n = 3; 15 cells/experiment). Data represent mean ± SD (ns, not significant; ***, P < 0.001; Student’s t test).