Cloning and functional expression of rNBC, an electrogenic Na+-HCO3/- cotransporter from rat kidney

Michael F Romero, Peying Fong, Urs V. Berger, Matthias A. Hediger, Walter F. Boron

Research output: Contribution to journalArticle

179 Citations (Scopus)

Abstract

We have recently cloned the renal electrogenic Na+-bicarbonate cotransporter of the salamander Ambystoma tigrinum (aNBC) (M. F. Romero, M. A. Hediger, E. L. Boulpaep, and W. F. Boron. FASEB J. 10: 89, 1996; and Nature 387: 409-413, 1997). Here we report the cloning of a mammalian homolog of aNBC, named rNBC for rat Na+-bicarbonate cotransporter. NBC constitutes the major route for HCO3/- reabsorption and assists in Na+ reabsorption across the basolateral membrane of the renal proximal tubule (PT). We used aNBC as a probe to screen a rat kidney cortex cDNA library in λgt10 and identified several clones. Each has an initiator Met and a large open- reading frame followed by a 3'-untranslated region of ~500 bp. The 7.5-kb mRNA for rNBC is present in kidney, liver, lung, brain, and heart. In situ hybridization with the rNBC probe in the rat kidney revealed staining in the S2 segment of PT. rNBC encodes a protein of 1,035 amino acids, with a predicted molecular mass of 116 kDa. Its deduced amino acid sequence is 86% identical to that of aNBC. Comparison of both the aNBC and rNBC sequences to the GenBank database reveals a low level of amino acid identity (~30%) to the AE family of Cl-/HCO3/- exchangers. Injection of rNBC cRNA into Xenopus oocytes leads to expression of an electrogenic Na+-HCO3/- cotransporter that is qualitatively similar to that of aNBC but at a much lower level. Placement of the rNBC cDNA into the context of a Xenopus expression vector produces a substantial increase in rNBC expression. Addition of 1.5% CO2/10 mM HCO3/- elicits a hyperpolarization of >50 mV and a rapid decrease of intracellular pH (pH(i)), followed by an increase in pH(i). Subsequent removal of Na+ in the presence of CO2/HCO3/- causes a depolarization of >50 mV and a concomitant decrease of phi. Thus rNBC is in the same newly identified family of Na+-linked HCO3/- transporters as is aNBC.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Renal Physiology
Volume274
Issue number2 43-2
StatePublished - Feb 1998
Externally publishedYes

Fingerprint

Organism Cloning
Bicarbonates
Xenopus
Kidney
Chloride-Bicarbonate Antiporters
Ambystoma
Amino Acids
Urodela
Kidney Cortex
Complementary RNA
Proximal Kidney Tubule
Nucleic Acid Databases
3' Untranslated Regions
Gene Library
Open Reading Frames
Oocytes
In Situ Hybridization
Amino Acid Sequence
Complementary DNA
Clone Cells

Keywords

  • Bicarbonate transport
  • Homology cloning
  • Intracellular pH
  • Rat sodium-bicarbonate cotransport
  • Sodium transport
  • Xenopus oocyte expression

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Cite this

Cloning and functional expression of rNBC, an electrogenic Na+-HCO3/- cotransporter from rat kidney. / Romero, Michael F; Fong, Peying; Berger, Urs V.; Hediger, Matthias A.; Boron, Walter F.

In: American Journal of Physiology - Renal Physiology, Vol. 274, No. 2 43-2, 02.1998.

Research output: Contribution to journalArticle

Romero, Michael F ; Fong, Peying ; Berger, Urs V. ; Hediger, Matthias A. ; Boron, Walter F. / Cloning and functional expression of rNBC, an electrogenic Na+-HCO3/- cotransporter from rat kidney. In: American Journal of Physiology - Renal Physiology. 1998 ; Vol. 274, No. 2 43-2.
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abstract = "We have recently cloned the renal electrogenic Na+-bicarbonate cotransporter of the salamander Ambystoma tigrinum (aNBC) (M. F. Romero, M. A. Hediger, E. L. Boulpaep, and W. F. Boron. FASEB J. 10: 89, 1996; and Nature 387: 409-413, 1997). Here we report the cloning of a mammalian homolog of aNBC, named rNBC for rat Na+-bicarbonate cotransporter. NBC constitutes the major route for HCO3/- reabsorption and assists in Na+ reabsorption across the basolateral membrane of the renal proximal tubule (PT). We used aNBC as a probe to screen a rat kidney cortex cDNA library in λgt10 and identified several clones. Each has an initiator Met and a large open- reading frame followed by a 3'-untranslated region of ~500 bp. The 7.5-kb mRNA for rNBC is present in kidney, liver, lung, brain, and heart. In situ hybridization with the rNBC probe in the rat kidney revealed staining in the S2 segment of PT. rNBC encodes a protein of 1,035 amino acids, with a predicted molecular mass of 116 kDa. Its deduced amino acid sequence is 86{\%} identical to that of aNBC. Comparison of both the aNBC and rNBC sequences to the GenBank database reveals a low level of amino acid identity (~30{\%}) to the AE family of Cl-/HCO3/- exchangers. Injection of rNBC cRNA into Xenopus oocytes leads to expression of an electrogenic Na+-HCO3/- cotransporter that is qualitatively similar to that of aNBC but at a much lower level. Placement of the rNBC cDNA into the context of a Xenopus expression vector produces a substantial increase in rNBC expression. Addition of 1.5{\%} CO2/10 mM HCO3/- elicits a hyperpolarization of >50 mV and a rapid decrease of intracellular pH (pH(i)), followed by an increase in pH(i). Subsequent removal of Na+ in the presence of CO2/HCO3/- causes a depolarization of >50 mV and a concomitant decrease of phi. Thus rNBC is in the same newly identified family of Na+-linked HCO3/- transporters as is aNBC.",
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AU - Hediger, Matthias A.

AU - Boron, Walter F.

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KW - Intracellular pH

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KW - Sodium transport

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