Expression of transfected human Na+/H+ exchanger (NHE-1) in the the basolateral membrane of opossum kidney cells.
Some epithelial cells have Na+/H+ exchanger (NHE) activity in both apical and basolateral membranes. Amiloride-sensitive NHE-1 is generally identified in the basolateral membrane. The renal cell line, OK7a, targets amiloride-resistant NHE predominantly to the apical membrane. It is controversial whether the transfected NHE-1 is targeted preferentially to the basolateral membrane in OK7a cells, when human NHE-1 is chronically expressed under control of constitutively active promoters. We tried to identify the membranes in which the transfected human NHE-1 could be detected following acute expression in OK7a cells. We have always observed small Na(+)-dependent pH recovery in the basolateral membrane in OK7a cells. It is, however, controversial whether or not OK7a cells express NHE activity in the basolateral membrane. We also characterized Na(+)-dependent pH recovery in the basolateral membrane. It was not inhibited by [4,4'diisothiocyanatostilbene-2,2'-disulfonic acid] (DIDS), [4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid] (SITS), or contralateral amiloride. Li+ but not K+, chol+, or NMG+ could replace Na+. These results are consistent with the presence of the NHE in the basolateral membrane. NHE activities were predominant in the apical membrane and those in both membranes were resistant to amiloride analogs. After stable transfection with human NHE-1 in a vector utilizing the metallothionein promoter, overnight induction with Zn(2+)increased the NHE activity and its sensitivity to amiloride only in the basolateral membrane in OK7a cells. We conclude that the transfected human NHE-1 is exclusively targeted to the basolateral membrane of OK7a cells during acute induction.
Takaichi
K
K
Fourth Department of Internal Medicine, University of Tokyo School of Medicine, Japan.
Miyajima
Y
Y
Hanai
J I
JI
Kurokawa
K
K
Fujita
T
T
Warnock
D G
DG
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000584:Amiloride; D000818:Animals; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002794:Choline / Q000494:pharmacology; D004232:Diuretics; D004847:Epithelial Cells / Q000737:chemistry* / Q000502:physiology; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000166:cytology*; D008082:Lipotropic Agents / Q000494:pharmacology; D008094:Lithium / Q000493:pharmacokinetics; D008565:Membrane Proteins / Q000235:genetics; D009893:Opossums; D011188:Potassium / Q000493:pharmacokinetics; D011401:Promoter Regions, Genetic / Q000502:physiology; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics*; D014162:Transfection
D004232:Diuretics; D008082:Lipotropic Agents; D008565:Membrane Proteins; D017923:Sodium-Hydrogen Exchangers; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000584:Amiloride; D008094:Lithium; D012964:Sodium; D002794:Choline; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D011188:Potassium