The European Eel NCCβ Encodes for an Electroneutral NaCl Cotransporter That Is Not Sensitive to Thiazide Diuretics

Document Type

Article

Publication Date

4-2014

Publication Title

FASEB Journal

DOI

10.1096/fasebj.28.1_supplement.1099.1

ISSN

1530-6860

Abstract

The renal thiazide-sensitive NaCl cotransporter (NCC) in mammalian nephron plays a key role in salt transport of the distal convoluted tubule and thus in modulation of arterial blood pressure and potassium secretion. Only one gene encoding NCC has been described in mammals. However, in the teleost European eel, two genes encoding putative isoforms of NCC were identified: NCCα, from kidney, and NCCβ from intestine. For the NCCα the identify degree with flounder, rat and human NCC is 70, 65, and 45%, respectively and for the NCCβ is 55, 55, and 43, respectively. No functional characterization of these variants has been reported. By RT-PCR we cloned the full-length cDNA of eel NCCβ isoform and inserted into the Xenopus expression vector pgh19. The insert contains 3437 bp, with an open reading frame of 3132, encoding a 1043 amino acid protein. Functional expression was assessed in Xenopus oocytes, microinjected with 10 ng of cRNA in vitro transcribed from NCCβ cDNA, by measuring the tracer 22Na+ uptake. Rat NCC was used as a control signal. Our results revealed that injection of oocytes with NCCβ cRNA induced the appearance of Na+ uptake mechanism that is fully dependent on extracellular Cl-, constituting a NaCl cotransporter. However, in strict contrast to rat NCC, eel NCCβ is not inhibited by thiazide type diuretics. In addition, it is also not stimulated by WNK3 kinase that is a powerful activator of mammalian NCC. We conclude that NCCβ encodes a non-thiazide sensitive NaCl cotransporter.

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