Figure 3

In vitro characterization of the NPT2c variants. (a) Na-dependent phosphate uptake: HEK expressing WT NPT2c or NPT2c variants (cells transfected with 0.5 μg of DNA and 1 μl of lipofectamine 2000 per well). 1 variant transport phosphate normally, 4 variants have a phosphate transport function slightly lowered et 8 have a strong decrease in phosphate transport. Quantitative data represent at least 9 independent experiments. Statistical analysis was performed using a non-parametric Kruskall–Wallis test followed by Dunn’s multiple comparison post-hoc test. ****p < 0.0001, simple line represents Kruskall–Wallis test. *p < 0.05, **p < 0.01, ****p < 0.0001 on NPT2c conditions represent Dunn’s multiple comparison post-hoc test compared to non- transfected cells. (b) We performed immunofluorescence on HEK cells transfected with transporting and non-transporting NPT2c variants (cells transfected with 0.5 μg of DNA and 1 μl of lipofectamine 2000 per well). All the transporting variants were correctly addressed at the plasmic membrane. Some of the non-transporting variants were not addressed at the plasmic membrane (e.g. c.1242C > G, p.Tyr414*), others were correctly addressed at the plasmic membrane (e.g. c.575C > T, p.Ser192Leu and c.1361A > G, p.Asn454Ser).