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. 2016 Jun 15;291(31):15940–15957. doi: 10.1074/jbc.M116.726935

FIGURE 7.

FIGURE 7.

SLC30A10N43H retains manganese efflux activity. A, HeLa cells were transfected with FLAG-tagged SLC30A10N43H. Two days after transfection, cultures were processed for immunofluorescence. SLC30A10 was detected using a monoclonal antibody against the FLAG epitope, and a polyclonal antibody against calnexin was used to demarcate the endoplasmic reticulum. P represents the Pearson's coefficient for colocalization between SLC30A10 and calnexin (mean ± S.E.; n = 13 cells). Scale bar, 10 μm. B, HeLa cells were cotransfected with FLAG-tagged SLC30A10N43H and HA-tagged DPP4. Two days after transfection, cultures were processed for immunofluorescence. SLC30A10 was detected using a polyclonal antibody against the FLAG tag. A monoclonal antibody against HA was used to detect DPP4. P represents the Pearson's coefficient for colocalization between SLC30A10 and DPP4 (mean ± S.E.; n = 15 cells). Scale bar, 10 μm. C, intracellular manganese levels were measured in HeLa cells transfected with control (denoted as Control), SLC30A10WT (WT), or SLC30A10N43H (N43H) constructs, exactly as described in Fig. 5 (mean ± S.E.; n = 3 for each construct; *, p < 0.05 for the difference between cells expressing SLC30A10WT and all other transfection conditions by one-way ANOVA and Dunnett's post hoc test; n.s. stands for “not significant” and denotes that the difference between WT and N43H groups was not statistically significant (p > 0.05)). D, cell viability was assessed in HeLa cells transfected with control (denoted as Control), SLC30A10WT (WT), or SLC30A10N43H (N43H) constructs after treatment with 0 or 2 mm manganese, exactly as described in Fig. 6E. For each transfection condition, viability at 0 mm manganese was set to 100 and used to calculate percent viability at 2 mm manganese (mean ± S.E.; n ≥ 5 for each construct; *, p < 0.05 for the difference between SLC30A10WT and all other transfection conditions by one-way ANOVA and Dunnett's post hoc test; n.s. stands for not significant and denotes that the difference between WT and N43H groups was not statistically significant (p > 0.05)).