(A) The localization of GFP WT-CLS and Myc-tagged cyclin E SWNQ-A without (left) and with (right) fusion to the PACT domain was analyzed by immunofluorescence in cells expressing the GFP-WT-CLS. Staining was performed with the indicated antibodies as described in Experimental Procedures: DAPI (blue), anti-GFP (green), and anti-Myc (red). Arrowheads mark centrosomes. Inset: enlarged image of the merged centrosomal area. Scale bars: 5 μM.
(B) DNA synthesis is restored in CLS-expressing cells by co-expression of PACT-fused Myc-cyclin E SWNQ-A. Cells were synchronized by double thymidine treatment, transfected with the indicated constructs, and assessed for BrdU incorporation at 1, 3, and 6 hr after release of G1-S arrest by thymidine washout. Error bars represent mean ± SD (n=5).