A common sequence variation of the CLCNKB gene strongly activates ClC-Kb chloride channel activity.
BACKGROUND
Tubular transepithelial reabsorption of chloride along the nephron is a major determinant of body salt and water homeostasis and blood pressure regulation. About 40% of the glomerulary filtered sodium chloride are reabsorbed in the distal nephrons. Vectorial transepithelial sodium chloride transport is critically dependent on the function of basolateral ClC-K type chloride channels there. Modulation of ClC-Kb chloride channel activity by polymorphic variations of the CLCNKB gene, thus, could form a molecular basis for salt sensitivity of blood pressure regulation. In this study we tested the effect of several polymorphic variants on ClC-Kb chloride channel activity.


METHODS
After heterologous expression in Xenopus oocytes, ClC-Kb channel activity and surface expression in presence of the ClC-K beta subunit barttin were determined by two-electrode voltage-clamp analysis, immunofluorescence, and ClC-Kb surface enzyme-linked immunosorbent assay (ELISA).


RESULTS
Chloride currents induced by the ClC-Kb variants L27R, G214A, I419V, T562M, and E578K were not significantly different from wild-type currents. The ClC-KbT481S variation, however, which showed a frequency of 20% in our control population, dramatically activated chloride conductance by a factor of 20. Activation of chloride currents was also observed after introducing homologous mutations in ClC-Ka and ClC-K1, but not in ClC-2 and ClC-5 chloride channels. ClC-Kb activation by the T481S mutation did not change intrinsic ion channel pore properties and did not require increased surface expression of ClC-KbT481S.


CONCLUSIONS
Genetic heterogeneity of ClC-Kb chloride channels correlates with functional heterogeneity, which assigns ClC-Kb to a set of genes potentially relevant for polygenic salt-sensitivity of blood pressure regulation.
Jeck
Nikola
N
Department of Pediatrics, Philipps University of Marburg, Germany.
Waldegger
Petra
P
Doroszewicz
Jolanta
J
Seyberth
Hannsjörg
H
Waldegger
Siegfried
S
D000818:Animals; D027321:Anion Transport Proteins / Q000235:genetics* / Q000378:metabolism*; D001794:Blood Pressure / Q000235:genetics; D018118:Chloride Channels / Q000235:genetics* / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D018740:Genetic Heterogeneity; D006801:Humans; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D009399:Nephrons / Q000378:metabolism; D018408:Patch-Clamp Techniques; D017354:Point Mutation; D012965:Sodium Chloride / Q000378:metabolism; D014982:Xenopus laevis
D027321:Anion Transport Proteins; C412733:CLCNKB protein, human; D018118:Chloride Channels; D002712:Chlorides; D008565:Membrane Proteins; D012965:Sodium Chloride