Cloning and characterization of novel human SLC4A8 gene products encoding Na+-driven Cl-/HCO3(-) exchanger variants NDCBE-A, -C, and -D

Physiol Genomics. 2008 Aug 15;34(3):265-76. doi: 10.1152/physiolgenomics.90259.2008. Epub 2008 Jun 24.

Abstract

The reported sequences of the human and mouse Na+-driven Cl-/HCO3(-) exchangers (NDCBEs) differ greatly in their extreme cytosolic COOH termini (Ct). In human NDCBE (NDCBE-B), a 17-amino acid (aa) sequence replaces 66 aa at the equivalent position in mouse NDCBE (NDCBE-A). We performed 5'- and 3'-rapid amplification of cDNA ends (RACE) on human brain cDNA, followed by PCR of full-length cDNAs to determine whether the human SLC4A8 gene was capable of producing the mouselike Ct sequence. Our study confirmed the presence in human cDNA of mouse NDCBE-like transcripts (human NDCBE-A) and also disclosed the existence of three further novel NDCBE transcripts that we have called NDCBE-C, NDCBE-D, and NDCBE-D'. The novel NDCBE-C/D/D' transcripts initiate at a novel "exon 0" positioned approximately 35 kb upstream of the first exon of NDCBE-A/B. NDCBE-C/D/D' protein products are predicted to be truncated by 54 aa in the cytosolic NH(2) terminus (Nt) compared with NDCBE-A/B. Our data, combined with a new in silico analysis of partial transcripts reported by others in the region of the human SLC4A8 gene, increase the known extent of the SLC4A8 gene by 49 kb, to 124 kb. A functional comparison of NDCBE-A/B/C/D expressed in Xenopus oocytes demonstrates that the Nt variation does not affect the basal functional expression of NDCBE, but those with the shorter Ct have a 25-50% reduced functional expression compared with those with the longer Ct. By comparison with an artificially truncated NDCBE that contains neither 17-aa nor 66-aa Ct cassette, we determined that the functional difference is unrelated to the 66-aa cassette of NDCBE-A/C, but is instead due to an inhibitory effect of the 17-aa cassette of NDCBE-B/D.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / pharmacology
  • Alternative Splicing / drug effects
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Biological Transport / drug effects
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Chloride-Bicarbonate Antiporters / chemistry
  • Chloride-Bicarbonate Antiporters / genetics*
  • Chloride-Bicarbonate Antiporters / metabolism
  • Chromosomes, Human
  • Cloning, Molecular
  • Exons / genetics
  • Humans
  • Hydrogen-Ion Concentration
  • Membrane Potentials / drug effects
  • Molecular Sequence Data
  • Mutant Proteins / metabolism
  • Nucleic Acid Amplification Techniques
  • Oocytes / cytology
  • Protein Isoforms / chemistry
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Sodium / metabolism
  • Sodium-Bicarbonate Symporters / chemistry
  • Sodium-Bicarbonate Symporters / genetics*
  • Sodium-Bicarbonate Symporters / metabolism
  • Solubility / drug effects
  • Xenopus

Substances

  • Chloride-Bicarbonate Antiporters
  • Mutant Proteins
  • Protein Isoforms
  • RNA, Messenger
  • SLC4A8 protein, human
  • Sodium-Bicarbonate Symporters
  • Sodium
  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid

Associated data

  • GENBANK/DQ063579
  • GENBANK/DQ975204
  • GENBANK/DQ996398
  • GENBANK/DQ996537