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Fig. 5 | Molecular Cancer

Fig. 5

From: MIEN1, a novel interactor of Annexin A2, promotes tumor cell migration by enhancing AnxA2 cell surface expression

Fig. 5

MIEN1 activates translocation of AnxA2 to plasma membrane to promote breast cancer cell migration. a Control and MIEN1 transfected MDA-MB231 cells were subjected to western blot analysis with anti-MIEN1 or anti-AnxA2 antibody. PGK served as a loading control. b Down-regulation of MIEN1 and AnxA2 reduces the abilities of MDA-MB231 breast cancer cells to migrate in vitro. Confluent MDA-MB231 monolayers transfected with control GFP, AnxA2 or/and MIEN1 were wounded using a pipet tip, 60 h after transfection. Following the wound formation, plates were incubated for 12 h and the wound closure areas were visualized on an Olympus Microscope (Carl Zeiss). Representative images were acquired at 0 and 12 h. c Quantification of cell migration was achieved using Image J software. Data are presented as percentages of the recovered scratch area relative to untreated control cells. Columns are the means five replicates from two independent experiments and bars are s.d. d MDA-MB231 and MDA-MB231 cells stably transfected with control GFP or GFP-MIEN1WT were subjected to western blot analysis with anti-phosho AnxA2, anti-AnxA2 and anti-MIEN1 antibodies. GAPDH served as a loading control. The quantification of the representative blots is the densitometric average of three independent experiments analyzed using ImageJ and normalized to GAPDH. e Control and MIEN1 transfected MDA-MB231 cells were subjected to western blot analysis with anti-phosho AnxA2, AnxA2, anti-MIEN1 and actin as loading control. f Representative images, as captured using a 60X oil immersion TIRF microscope, of MDA-MB231 and MDA-MB231 cells stably transfected with GFP-MIEN1WT grown to sub-confluence on coverslips, fixed with PFA, unpermeabilized, and processed for TIRF microscopy with the specific antibodies. At least three independent fields were analyzed for confirmation of the staining pattern represented. Scale bar denotes 20 μm

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