Figure 7. Overexpression of survivin rescues the defects in CUL7-depleted cells.

(A) U2OS cells were transfected with empty vector or different amounts of plasmids expressing survivin. 24 hours after transfection, cells were transfected again with scrambled or targeting CUL7 targeting siRNA oligos. 48 hours later, the cells were photographed and a representative phase contrast view of each sample is shown (Top); the expression of individual proteins was confirmed by immunoblotting (left). The cells were then collected by trypsinization, stained with trypan blue and viable cells were counted and plotted (right).

(B) U2OS cells were transfected with empty vector or Flag-survivin for 24 hours and then transfected again with indicated siRNA oligos. 48 hours later, the cells were lysed and analyzed by immunoblotting with indicated antibodies, and densitometry analysis with ImageJ software was performed to obtain acetyl-α-tubulin band intensity relative to α-actin (right; *p < 0.001).

(C) U2OS cells stably expressing pEGFP-α-tubulin were transiently transfected as indicated and cultured in nutrient medium for 48 hours before recording images. A representative of confocal Z-stack image (6 slides) is shown. The scale bar is 10 µm.

(D) A schematic model illustrating the 3M–CUL9-survivin pathway.