GRASP depletion–mediated Golgi destruction decreases cell adhesion and migration via the reduction of a5ß1 integrin

Ahat, Erpan; Xiang, Yi; Zhang, Xiaoyan; Bekier, Michael E.; Wang, Yanzhuang

The wound-healing assay used here was performed using the protocol described by Jun-Lin Guan and colleagues (Liang et al., 2007) . MDA-MB-231 or HeLa cells were transfected with the indicated plasmids; and at 48 h after transfection, the cells were replated in 12-well plates at 80% confluence in complete growth medium. Twenty-four hours later, after the cells became confluent and formed a monolayer, the medium was replaced with serumfree DMEM. Twenty-four hours afterward, the cell monolayer was scraped in a straight line to create a "scratch" with a 200-µl pipette tip. The cells were washed once with DMEM, and images were taken using an inverted bright-field microscope with a 4× lens and an IMAGING ERTIGA 1300R camera. The cells were then allowed to grow in complete growth medium for 20 h, and images of the same area were taken. Images were analyzed with ImageJ software (National Institutes of Health) or WimScratch: Wound Healing Assay Image Analysis Solution (Release 4.0; available from www.wimasis.com/en/products/9/WimScratch) (Jameson et al., 2013) .