Integrin α6β4 binds to and promotes secretion of laminin-5 in Panc1 cells. (A–C) Panc1 clones 3D7 (A; high α6β4) and 2G6 (B; low α6β4) were plated on collagen I coated coverslips and allowed to adhere under normal culture conditions for 4 hours. Cells were fixed and immunostained for integrin β4 subunit (red), laminin-5 (γ2 subunit; green), or f-actin (blue) as described in the Methods section. Cells were imaged by TIRF microscopy using the same exposure times and settings. Pearson’s coefficient for colocalization between integrin β4 and laminin-5 were 0.8 for Panc1-3D7 (A) and 0.011 for Panc1-2G6 (B). These values are representative for the 30 cells analyzed for each cell line. (C) Represents a rare β4 expressing cell in the Panc-2G6 cell population. (D) Western blot analysis of whole cell extracts from MDA-MB-231 (positive control), Panc1-2G6 and Panc1-3D7 cells for integrin β4, laminin-5 and actin (loading control).