Differential expression of AE1 in renal HCO3-secreting and -reabsorbing intercalated cells.
The cortical collecting duct of the kidney contains two types of intercalated cells that transport HCO3 in opposite directions. HCO3 reabsorption takes place in the alpha-type intercalated cells, which express a Cl/HCO3 exchanger on the basolateral membrane. This exchanger is the product of the anion exchanger 1 (AE1) or band 3 gene. HCO3 secretion occurs in the beta-intercalated cells, which have a Cl/HCO3 exchanger on the apical membrane. Based on studies in an immortalized cell line, recently it was proposed that the apical anion exchanger of beta-intercalated cells is also AE1 (van Adelsberg, J. S., Edwards, J. C., and Al-Awqati, Q. (1993) J. Biol. Chem. 268, 11283-11289). In the present study we reinvestigated this issue by determining the distribution of AE1 mRNA and protein in the two intercalated cell types using cells freshly isolated from the native epithelium. Using quantitative reverse transcriptase polymerase chain reaction, we found that alpha-intercalated cells, isolated from rabbit kidney by fluorescence-activated cell sorting, have high levels of AE1 mRNA, whereas beta-intercalated cells express very low levels. The ratio of AE1 mRNA levels in alpha- versus beta-intercalated cells averaged 10.1 +/- 2.6. In addition, metabolic acidosis increased the levels of AE1 mRNA by 3-5-fold in cortical collecting duct cells. This difference was confirmed by Northern blotting. Western blotting using an antibody against rabbit AE1 revealed a major immunoreactive product with a molecular weight of approximately 110 kDa in cortical collecting duct cells. Deglycosylation reduced the size of the immunoreactive product to approximately 90 kDa, which is compatible with the presence of a truncated form of AE1. Metabolic acidosis increased the intensity of the AE1 immunoreactive band. The level of AE1-immunoreactive protein was significantly higher in alpha-intercalated cells than in beta-intercalated cells. In aggregate, these data provide evidence for the differential expression of AE1 in HCO3-reabsorbing versus HCO3-secreting renal intercalated cells both at the mRNA and at the protein level. These results give no support to the concept that AE1 functions both as a basolateral and an apical anion exchanger in cortical collecting duct cells.
Fejes-Tóth
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G
Department of Physiology, Dartmouth Medical School, Lebanon, New Hampshire 03756.
Chen
W R
WR
Rusvai
E
E
Moser
T
T
Náray-Fejes-Tóth
A
A
D000138:Acidosis / Q000378:metabolism; D000471:Alkalosis / Q000378:metabolism; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D001639:Bicarbonates / Q000378:metabolism*; D001692:Biological Transport; D002469:Cell Separation; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D007672:Kidney Cortex / Q000166:cytology / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D014018:Tissue Distribution
D001457:Anion Exchange Protein 1, Erythrocyte; D001639:Bicarbonates; D012333:RNA, Messenger