Figure 3.

Hakai regulates Annexin A2 expression levels: (a) HEK293T cells were transiently transfected with pcDNA-Flag-Hakai (4 µg), pBSSR-HA-ubiquitin (3 µg) and pSG-v-Src (3 µg) plasmids for 48 h. Cells were harvested and the levels of endogenous Annexin A2 levels and Hsp90 were determined by western blot analysis (left panel) and quantified by densitometry (right panel). The effect of the indicated transfected siRNA-Hakai on Annexin A2 and Hsp90 levels were tested in 293T cells (b) and HCT116 (c). Whole-cell lysates were subjected to western-blotting 72 h after transfection (top) and protein expression was quantified by densitometry (bottom) using GAPDH as loading control for normalization. Relative quantification of Annexin A2 expression levels was graphically represented as Mean ± SEM for two independent experiments for panel a and three for panel b and c (* p < 0.05, ** p < 0.01).