ClC-5: role in endocytosis in the proximal tubule.
The proper functioning of the Cl(-) channel, ClC-5, is essential for the uptake of low molecular mass proteins through receptor-mediated endocytosis in the proximal tubule. Dent's disease patients with mutant ClC-5 channels and ClC-5 knockout (KO) mice both have low molecular mass proteinuria. To further understand the function of ClC-5, endocytosis was studied in LLC-PK(1) cells and primary cultures of proximal tubule cells from wild-type (WT) and ClC-5 KO kidneys. Endocytosis in the proximal tubule cells from KO mice was reduced compared with that in WT animals. Endocytosis in WT but not in KO cells was inhibited by bafilomycin A-1 and Cl(-) depletion, whereas endocytosis in both WT and KO cells was inhibited by the NHE3 blocker, S3226. Infection with adenovirus containing WT ClC-5 rescued receptor-mediated endocytosis in KO cells, whereas infection with any of the three disease-causing mutants, myc-W22G-ClC-5, myc-S520P-ClC-5, or myc-R704X-ClC-5, did not. WT and the three mutants all trafficked to the apical surface, as assessed by surface biotinylation. WT-ClC-5 and the W22G mutant were internalized similarly, whereas neither the S520P nor the R704X mutants was. These data indicate that ClC-5 is important for Cl(-) and proton pump-mediated endocytosis. However, not all receptor-mediated endocytosis in the proximal tubule is dependent on ClC-5. There is a significant fraction that can be inhibited by an NHE3 blocker. Our data from the mutants suggest that defective targeting and trafficking of mutant ClC-5 to the endosomes are a major determinant in the lack of normal endocytosis in Dent's disease.
Wang
Yinghong
Y
Dept. of Physiology, WBSB Rm. 208, The Johns Hopkins Univ. School of Medicine, 725 N. Wolfe St., Baltimore, MD 21205, USA.
Cai
Hui
H
Cebotaru
Liudmila
L
Hryciw
Deanne H
DH
Weinman
Edward J
EJ
Donowitz
Mark
M
Guggino
Sandra E
SE
Guggino
William B
WB
D000818:Animals; D015153:Blotting, Western; D018118:Chloride Channels / Q000235:genetics* / Q000502:physiology; D003911:Dextrans / Q000494:pharmacology; D004705:Endocytosis / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D007687:Kidney Tubules, Proximal / Q000502:physiology*; D018374:LLC-PK1 Cells; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation / Q000235:genetics / Q000502:physiology; D010957:Plasmids / Q000235:genetics; D017494:Proton Pumps / Q000502:physiology; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000502:physiology; D013552:Swine; D014162:Transfection
C097503:CLC-5 chloride channel; D018118:Chloride Channels; D003911:Dextrans; D017494:Proton Pumps; C000613386:Slc9a3 protein, mouse; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers