Fig. 3.

Mps1 localization to the centrosome is critical for its centrosomal functions. (A) Representative images of asynchronously growing HeLa cells transfected with indicated constructs and stained for BrdU to identify S-phase cells, showing BrdU (blue), GFP (green), centrioles (Cetn2, red), and centrosomes (γ-tub, magenta). (B) Percentage of cells positive for BrdU and GFP with two or fewer centrioles, as judged by Cetn2 foci; mean ± SD of n = 3 independent experiments of n = 100 cells for each experiment. (C) Immunoblot analysis of HEK293 cells transfected with either siControl (siRNA against Lamin A/C) or siMps1 for 24 h before being transfected with indicated siRNA-resistant constructs for 48 h. For the final 24 h, cells were arrested in S-phase with HU. Cells were lysed, separated via SDS/PAGE, and blotted with anti-GFP to assess transfected protein level, and anti-pT686 to assess Mps1 activity, with alpha tubulin (α-tub) serving as a loading control. An asterisk signifies a nonspecific band. (Scale bars, 5 µm.)