Department of Internal Medicine, Faculty of Medicine, University of Tokyo, 7-3-1 Bunkyo-ku, Hongo, Tokyo, 113-0033, Japan.
Pflugers Arch. 2011 Feb;461(2):249-59. doi: 10.1007/s00424-010-0918-x. Epub 2011 Jan 14.
The electrogenic Na(+)-HCO(3)(-) cotransporter NBCe1 encoded by SLC4A4 plays essential roles in the regulation of intracellular/extracellular pH. Homozygous mutations in NBCe1 cause proximal renal tubular acidosis associated with ocular abnormalities. In the present study, we tried to perform functional characterization of the four nonsynonymous single nucleotide polymorphisms (SNPs), E122G, S356Y, K558R, and N640I in NBCe1A. Functional analysis in Xenopus oocytes revealed that while the K558R variant had a significantly reduced transport activity corresponding to 47% of the wild-type activity, the remaining variants E122G, S356Y, and N640I did not change the NBCe1A activity. Apparent Na(+) affinity of K558R was not different from that of wild-type NBCe1A. Immunohistological analyses in HEK293 cells and MDCK cells indicated that none of these SNPs changed the trafficking behaviors of NBCe1A. Functional analysis in HEK293 cells also revealed that only the K558R variant had a reduced transport activity, corresponding to 41-47% of the wild-type activity. From these results, we conclude that among four SNPs, only the K558R variant, which is predicted to lie in transmembrane segment 5, significantly reduces the NBCe1A activity without changing the trafficking behavior or the apparent extracellular Na(+) affinity.
SLC4A4 编码的生电性 Na(+)-HCO(3)(-)共转运蛋白 NBCe1 在调节细胞内外 pH 值方面发挥着重要作用。NBCe1 的纯合突变会导致与眼部异常相关的近端肾小管酸中毒。在本研究中，我们试图对 NBCe1A 中的四个非同义单核苷酸多态性 (SNP)，即 E122G、S356Y、K558R 和 N640I 进行功能特征分析。在非洲爪蟾卵母细胞中的功能分析表明，虽然 K558R 变体的转运活性显著降低，仅为野生型活性的 47%，但其余变体 E122G、S356Y 和 N640I 并未改变 NBCe1A 的活性。K558R 的表观 Na(+)亲和力与野生型 NBCe1A 没有差异。在 HEK293 细胞和 MDCK 细胞中的免疫组织化学分析表明，这些 SNP 均未改变 NBCe1A 的运输行为。在 HEK293 细胞中的功能分析还表明，只有 K558R 变体的转运活性显著降低，仅为野生型活性的 41-47%。根据这些结果，我们得出结论，在这四个 SNP 中，只有预测位于跨膜片段 5 中的 K558R 变体显著降低了 NBCe1A 的活性，而不改变运输行为或表观细胞外 Na(+)亲和力。
