Selectively amplified expression of an isoform of the vacuolar H(+)-ATPase 56-kilodalton subunit in renal intercalated cells.
The intercalated cells of the kidney collecting duct are specialized for physiologically regulated proton transport. In these cells, a vacuolar H(+)-ATPase is expressed at enormous levels in a polarized distribution on the plasma membrane, enabling it to serve in transepithelial H+ transport. In contrast, in most eukaryotic cells, vacuolar H(+)-ATPases reside principally in intracellular compartments to effect vacuolar acidification. To investigate the basis for the selective amplification of the proton pump in intercalated cells, we isolated and sequenced cDNA clones for two isoforms of the approximately 56-kDa subunit of the H(+)-ATPase and examined their expression in various tissues. The predicted amino acid sequence of the isoforms was highly conserved in the internal region but diverged in the amino and carboxyl termini. mRNA hybridization to a cDNA probe for one isoform (the "kidney" isoform) was detected only in kidney cortex and medulla, whereas mRNA hybridization to the other isoform of the approximately 56-kDa subunit and to the H(+)-ATPase 31-kDa subunit was found in the kidney and other tissues. Immunocytochemistry of rat kidney with an antibody specific to the kidney isoform revealed intense staining only in the intercalated cells. Staining was absent from proximal tubule and thick ascending limb, where H(+)-ATPase was detected with a monoclonal antibody to the 31-kDa subunit of the H(+)-ATPase. This example of specific amplification of an isoform of one subunit of the vacuolar H(+)-ATPase being limited to a specific cell type suggests that the selective expression of the kidney isoform of the approximately 56-kDa subunit may confer the capacity for amplification and other specialized functions of the vacuolar H(+)-ATPase in the renal intercalated cell.
Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110.
R D
RD
Nelson
X L
XL
Guo
K
K
Masood
D
D
Brown
M
M
Kalkbrenner
S
S
Gluck
D000328:Adult; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000201:enzymology; D002417:Cattle; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000235:genetics*; D007668:Kidney / Q000201:enzymology*; D007672:Kidney Cortex / Q000201:enzymology; D007679:Kidney Medulla / Q000201:enzymology; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016376:Oligonucleotides, Antisense; D009928:Organ Specificity; D016133:Polymerase Chain Reaction / Q000379:methods; D006180:Proton-Translocating ATPases / Q000235:genetics*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D014617:Vacuoles / Q000201:enzymology
D007527:Isoenzymes; D046911:Macromolecular Substances; D016376:Oligonucleotides, Antisense; D012313:RNA; D004247:DNA; D006180:Proton-Translocating ATPases