Barttin increases surface expression and changes current properties of ClC-K channels

Abstract

The term Bartter syndrome encompasses a heterogeneous group of autosomal recessive salt-losing nephropathies that are caused by disturbed transepithelial sodium chloride reabsorption in the distal nephron. Mutations have been identified in the NKCC2 (Na(+)-K(+)-2Cl(-)) cotransporter and ROMK potassium channel, which cooperate in the process of apical sodium chloride uptake, and ClC-Kb chloride channels, which mediate basolateral chloride release. Recently, mutations in barttin, a protein not related to any known ion transporter or channel, were described in BSND, a variant of Bartter syndrome associated with sensorineural deafness. Here we show that barttin functions as an activator of ClC-K chloride channels. Expression of barttin together with ClC-K in Xenopus oocytes increased ClC-K current amplitude, changed ClC-K biophysical properties, and enhanced ClC-K abundance in the cell membrane. Co-immunoprecipitation revealed a direct interaction of barttin with ClC-K. We performed in situ hybridization on rat kidney slices and RT-PCR analysis on microdissected nephron segments to prove co-expression of barttin, ClC-K1 and ClC-K2 along the distal nephron. Functional analysis of BSND-associated point mutations revealed impaired ClC-K activation by barttin. The results demonstrate regulation of a CLC chloride channel by an accessory protein and indicate that ClC-K activation by barttin is required for adequate tubular salt reabsorption.

Identification of ClC-2 and CIC-K2 Chloride Channels in Cultured Rat Type IV Spiral Ligament Fibrocytes

“…8D). These electrophysiological characteristics closely resemble those of ClC-Kb channels (human homolog of ClC-K2) when expressed in Xenopus oocytes with the exception of pH, which has an opposite effect (Estevez et al 2001;Waldegger et al 2002).…”

Carboxyl-terminal Truncations of ClC-Kb Abolish Channel Activation by Barttin Via Modified Common Gating and Trafficking

“…However, in contrast to hClC-Ka and hClC-Kb, rClC-K1 can form functional channels also in the absence of barttin (4,6,7). This property permits studying the effects of barttin on rClC-K1 gating and allowed identification of mechanisms underlying the functional CLC-K modification by barttin (27,45).…”

“…Although hClC-Ka and hClC-Kb expression shows significant overlap, it is assumed that these two chloride channels exert distinct cellular functions. hClC-Ka participates in the transepithelial sodium chloride reuptake along the thin ascending limb of Henle facilitating water reabsorption in the adjacent thin descending limb of Henle whereas hClC-Kb predominantly mediates the basolateral chloride efflux necessary for salt absorption in the thick ascending limb of Henle (7)(8)(9).…”

Loss-of-function DNA sequence variant in the CLCNKA chloride channel implicates the cardio-renal axis in interindividual heart failure risk variation

“…We examined the functional implications of this gene variant by comparative analysis of recombinantly expressed Arg83 and Gly83 ClC-K a channels in tsA201cells (21). Because barttin is necessary for ClC-K functional expression (22,23), inward and outward currents in cells expressing ClC-K a Arg83 or Gly83 alone or barttin alone were no different from nontransfected or YFP-expressing cells (Fig. 3A).…”