Am J Physiol Cell Physiol 2007 Jun 01;2926:C2032-45. doi: 10.1152/ajpcell.00544.2006.

Cloning and characterization of an electrogenic Na/HCO3- cotransporter from the squid giant fiber lobe.

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The squid giant axon is a classic model system for understanding both excitable membranes and ion transport. To date, a Na(+)-driven Cl-HCO(3)(-) exchanger, sqNDCBE--related to the SLC4 superfamily and cloned from giant fiber lobe cDNA--is the only HCO(3)(-)-transporting protein cloned and characterized from a squid. The goal of our study was to clone and characterize another SLC4-like cDNA. We used degenerate PCR to obtain a partial cDNA clone (squid fiber clone 3, SF3), which we extended in both the 5' and 3' directions to obtain the full-length open-reading frame. The predicted amino-acid sequence of SF3 is similar to sqNDCBE, and a phylogenetic analysis of the membrane domains indicates that SF3 clusters with electroneutral Na(+)-coupled SLC4 transporters. However, when we measure pH(i) and membrane potential--or use two-electrode voltage clamping to measure currents--on Xenopus oocytes expressing SF3, the oocytes exhibit the characteristics of an electrogenic Na/HCO(3)(-) cotransporter, NBCe. That is, exposure to extracellular CO(2)/HCO(3)(-) not only causes a fall in pH(i), followed by a robust recovery, but also causes a rapid hyperpolarization. The current-voltage relationship is also characteristic of an electrogenic NBC. The pH(i) recovery and current require HCO(3)(-) and Na(+), and are blocked by DIDS. Furthermore, neither K(+) nor Li(+) can fully replace Na(+) in supporting the pH(i) recovery. Extracellular Cl(-) is not necessary for the transporter to operate. Therefore, SF3 is an NBCe, representing the first NBCe characterized from an invertebrate.

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