Novel CLCNKB mutations causing Bartter syndrome affect channel surface expression.
2013
Mutations in the
CLCNKBgene encoding the ClC-Kb Cl− channel cause
Bartter syndrome, which is a salt-losing renal
tubulopathy. Here, we investigate the functional consequences of seven mutations. When expressed in Xenopus laevis oocytes, four
mutantscarried no current (c.736G>C, p.Gly246Arg; c.1271G>A, p.Gly424Glu; c.1313G>A, p.Arg438His; c.1316T>C, p.Leu439Pro), whereas others displayed a 30%–60% reduction in conductance as compared with wild-type ClC-Kb (c.242T>C, p.Leu81Pro; c.274C>T, p.Arg92Trp; c.1052G>C, p.Arg351Pro). Anion selectivity and sensitivity to external Ca2+ and H+, typical of the ClC-Kb channel, were not modified in the partially active
mutants. In oocytes, we found that all the mutations reduced surface expression with a profile similar to that observed for currents. In HEK293 cells, the currents in the
mutantshad similar profiles to those obtained in oocytes, except for p.Leu81Pro, which produced no current. Furthermore, p.Arg92Trp and p.Arg351Pro mutations did not modify the unit-conductance of closely related ClC-K1. Western blot analysis in HEK293 cells showed that ClC-Kb protein abundance was lower for the nonconducting
mutantsbut similar to wild-type for other
mutants. Overall, two classes of
mutantscan be distinguished: nonconducting
mutantsassociated with low total protein expression, and partially conducting
mutantswith unaltered channel properties and ClC-Kb protein abundance.
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