Zinc accumulation upon ZnT2 overexpression decreases vesicular pH.

Error bars represent S.D. of at least 3 independent experiments. A) MCF-7 cells tranfected with ZnT2-Ruby were either incubated in growth medium, growth medium containing 75 μM ZnSO4, or growth medium supplemened with 5 μM TPEN and were analyzed for FluoZin3 fluorescence levels. The mean fluorescence levels were calculated only for live cells overexpressing ZnT2-Ruby. Asterisks indicate that the values obtained are significantly different from control cells in growth medium (p.05). B) LysoTracker Red fluorescence was determined in cells transfected with YFP empty vector (gray) or ZnT2-YFP (black) in various ZnSO4 concentrations. The median LysoTracker Red fluorescence level of the transfected cell population was divided by the value of the untransfected cells in the same experiment to calculate the ratio. Asterisks indicate that the values obtained are significantly different from control cells in growth medium without any additional ZnSO4 (p.05). C) Mean LysoTracker Red fluorescence was determined in cells transfected with YFP empty vector (gray) or ZnT2-YFP (black) incubated in growth medium or with the addition of 75 μM ZnSO4 or 5 μM TPEN. Asterisks indicate that the values obtained are significantly different from control cells in growth medium (p.05). D) Mean Lyso-pHluorin fluorescence levels were determined in cells transfected with ZnT2-Ruby incubated in growth medium or with additional 75 μM ZnSO4 or 5 μM TPEN. Asterisks indicate that the values obtained are significantly different from control cells in growth medium (p.05).