D405E and I803M SNVs of NBCe1 Show Reduced Cell-Surface Expression and Transport Activity

Background: The electrogenic Na+/HCO3- cotransporter NBCe1 is located in proximal renal tubule and regulate pH homeostasis. NBCe1 homozygous mutation causes many symptoms not only proximal renal tubular acidosis (RTA), but also extrarenal phenotype (migraine, mental retardation, mineral bone disorder and ocular abnormalities). Methods: We identified D405E and I803M missense mutations in NBCe1 variant A (kidney type) from the NCBI database. We performed the analysis of cellular localization and cell protein expression using confocal microscopy and western blotting. We also measured transport activity by using two-electrode voltage clamp method. Results: Immunofluorescence analysis with confocal microscopy revealed that the D405E and I803M variants were mainly localized in the cell-surface in HEK293 cells and MDCK cells. Western blotting in HEK293 cells confirmed that the D405E and I803M variants were significantly reduced protein expression level (35% in D405E and 53% in I803M) compared with WT, respectively. Moreover, electrophysiological analysis revealed that D405E and I803M variants were significantly reduced transport activity. Conclusion: These data suggest that D405E and I803M variants significantly reduced cell-expression level and affected transport activity.