Figure 6
Over-expression of ZIP1-Myc in RWPE2. RWPE2 was stably transfected with an empty vector or a ZIP1-myc expressing plasmid. a. Expression of the ZIP1-Myc mRNA in two stably transfected RWPE2 cell lines. Vector or ZIP-Myc transfected (Clone A and Clone B) RWPE2 were seeded at ~50% confluence and 0 or 25 μM ZnSO4 was added into the medium after 24 hours incubation. Cells were then incubated at 37°C for another 24 hours and total RNA was isolated. The endogenous ZIP1 as well as ZIP1-Myc mRNA were measured by quantitative RT-PCR with a TaqMan probe specific for ZIP1. The expression of ZIP1 mRNA was normalized to the β-actin mRNA expression. Values are the means ± SD of three experiments. b. Expression and localization of ZIP1-Myc in stably transfected RWPE2. Vector and ZIP1-Myc transfected RWPE2 were seeded at ~25% confluence and cultured at 37°C for 48 hours. ZIP1-Myc was detected by a mouse anti-Myc antibody (panel B). Arrow indicates the plasma membrane localization of ZIP1-Myc. The Myc staining of RWPE2 stably transfected with the vector control is shown in panel A. c. Expression of ZNT1 mRNA in stably transfected RWPE2. Vector or ZIP1-Myc expressing (Clone A and Clone B) RWPE2 were seeded at ~50% confluence and 0 or 25 μM ZnSO4 was added into the medium after 24 hours incubation. Cells were then incubated at 37°C for another 24 hours and total RNA was isolated. The endogenous ZNT1 mRNA was measured by quantitative RT-PCR with a TaqMan probe specific for ZNT1. The expression of ZNT1 was normalized to the β-actin expression. Values are the means ± SD of three experiments.