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Cloning of a unique electrogenic bicarbonate transporter from the squid giant fiber lobe
Author(s) -
Piermarini Peter M,
Choi Inyeong,
Boron Walter F
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a842
Subject(s) - squid giant axon , cotransporter , biophysics , chemistry , amino acid , reversal potential , biology , biochemistry , microbiology and biotechnology , membrane potential , sodium , patch clamp , receptor , organic chemistry
The squid ( Loligo pealei ) giant axon is a well‐studied model system for intracellular pH (pH i ) regulation, but the molecular identities of the proteins responsible for HCO 3 transport in this system are still unclear. To date, only one HCO 3 transporter—an electroneutral Na‐driven Cl/HCO 3 exchanger (sqNDCBE), related to the SLC4 family—has been cloned and characterized from squid giant fiber lobe (GFL) cDNA. Our goal was to identify and characterize another. Using PCR with degenerate primers, followed by 5′ and 3′ RACE, we identified a full‐length clone (SF3) that consists of 1,162 amino acids with 50% identity to sqNDCBE, 46–48% identity to human electroneutral NBCs, and 40–44% identity to human electrogenic NBCs. Phylogenetic analysis of the SF3 amino acid sequence reveals that SF3 clusters with sqNDCBE and other electroneutral NBCs, suggesting that SF3 is electroneutral. However, when Xenopus oocytes were injected with SF3 cRNA and exposed to a buffer containing 5% CO 2 /33 mM HCO 3 , the oocyte membrane potential showed a large, rapid hyperpolarization, followed by a slow relaxation, indicating that SF3 is electrogenic. Two‐electrode voltage clamping and pH i measurements demonstrate that the current and pH i recovery via SF3 depend on extracellular Na + but not Cl − , and are inhibited by DIDS. In conclusion, we have cloned from squid GFL cDNA an electrogenic Na‐dependent HCO 3 cotransporter (sqNBCe), which to our knowledge is the first electrogenic NBC characterized from an invertebrate.