A previously uncharacterized R881S variant of transporter NBCe1 exhibits intracellular retention and virtually no...

Mutations in the Na⁺/[Formula: see text] cotransporter NBCe1 (SLC4A4) cause proximal renal tubular acidosis (pRTA) and extrarenal symptoms including glaucoma, band keratopathy, migraine, growth disorder, and abnormal tooth enamel. From the NCBI dbSNP database, we recently identified a previously uncharacterized single-nucleotide variant (SNV), R881S, in NBCe1, located in hydrophilic helix 4. R881S NBCe1-A showed intracellular retention in human embryonic kidney 293 (HEK293) cells, and its plasma membrane expression was profoundly reduced in polarized Madin-Darby carine kidney (MDCK) cells. Unlike wild-type (WT) and R881C NBCe1-A (which causes pRTA), Western blot analysis demonstrated that the R881S NBCe1-A showed a single, low-molecular-weight signal above 100 kDa. Deglycosylation study showed that R881S NBCe1-A was scarcely deglycosylated by PNGase F. Coimmunoprecipitation study demonstrated that wild-type and R881S NBCe1-A did not form a heterodimer. Moreover, functional analysis using Xenopus oocytes revealed that the R881S variant had markedly reduced transport activity compared with wild-type NBCe1-A. Thus, R881S NBCe1-A shows no detectable transport activity, likely due to defective trafficking and reduced glycosylation. In contrast to the R881C mutant, however, R881S NBCe1-A fails to form dimers with wild-type NBCe1-A, suggesting the absence of a dominant-negative effect and underscoring the need for functional characterization of reported genetic variants.NEW & NOTEWORTHY Electrogenic Na⁺/[Formula: see text] cotransporter 1-A (NBCe1-A) in the proximal tubules regulates acid/base balance and fluid volume homeostasis. From the NCBI dbSNP database, we identified R881S NBCe1-A, which lacks glycosylation, cell-surface expression, and transport activity. We also found that the R881S variant does not form heterodimers with wild type, likely due to a reduced opportunity to interact with the wild-type protein at the plasma membrane.