FIGURE 6.
Streptavidin-bound biotin-bafilomycin is membrane-impermeable and inhibits in vitro invasion and migration of MB231 cells. A, vacuolar membranes were isolated from the yeast strain YPH500, and ATP-dependent proton transport activity was measured as described under “Experimental Procedures.” The values are the means of two independent experiments set relative to vacuoles treated with DMSO alone. B and C, MB231 cells were treated in the presence or absence of the V-ATPase specific inhibitor concanamycin A (ConA) or streptavidin-bound, biotin-conjugated bafilomycin and then plated on MatrigelTM-coated FluoroBlokTM inserts (B, invasion) or uncoated FluoroBlokTM inserts (C, migration) and allowed to migrate toward a chemoattractant on the trans-side of the well for 8 h. The cells were then stained with calcein AM, and the number of cells that had invaded or migrated to the trans-side was counted, with three wells analyzed per sample and an average of 10 images analyzed per well. The values for both assays are the means of at least three independent experiments expressed relative to cells treated with DMSO and streptavidin alone. All error bars indicate S.E. *, p < 0.05 compared with cells treated with DMSO and streptavidin alone.