Fig. 2.

mZip14 localizes to the plasma membrane of Sf9 cells and increases iron and zinc accumulation. (A) Sf9 cells were infected with wild-type baculovirus (control) or baculovirus containing the cDNA for mZip14. After 64 h, total cell lysates (15 μg of protein per lane) were analyzed by Western blotting as above. Lanes represent three independent experiments. (B and C) The same cells were analyzed by immunofluorescence microscopy. Cells were fixed and incubated with an anti-mZip14 antibody followed by goat anti-rabbit IgG-Alexa Fluor 594. Cells were counterstained with DAPI to visualize nuclei. (D) Iron accumulation measured with ⁵⁹Fe in medium containing 2 μM FeCl₂. (E) Zinc accumulation measured with ⁶⁵Zn in medium containing 2 μM ZnCl₂. (F) mZip14-dependent iron accumulation measured with ⁵⁹Fe in medium containing 2 μM FeCl₂ and 200 μM ZnCl₂. Cells were incubated at 27°C for 15 min in uptake medium. Values are the mean ± SD of three separate experiments, each with n = 3. ∗, P < 0.05 compared with control.