Dent's disease is an hereditary renal tubular disorder characterized by low-molecular-weight (LMW)
proteinuria,
hypercalciuria and
nephrolithiasis. The disease is due to mutations of CLC-5, a member of the family of voltage-gated CLC
chloride channels. CLC-5 is distributed in cells lining the proximal tubule (PT) of the kidney, where it co-localizes with
albumin-containing endocytic vesicles that form part of the receptor-mediated endocytic pathway that mediates the reabsorption of low-molecular-weight (LMW)
proteins filtered at the glomerular level. Since progression along the endocytic apparatus requires endosomal acidification, it has been suggested that dysfunction of CLC-5 in endosomes may lead to inefficient reabsorption of LMW
proteins and dysfunction of PT cells. Investigations conducted in a CLC-5 knockout (KO) mouse model harbouring all the characteristic renal tubular defects of
Dent's disease showed a severe impairment of endocytosis by PT cells, such that the endocytic tracer
peroxidase was poorly transferred into early endocytic vesicles. These data demonstrated that an impairment of receptor-mediated endocytosis in PT cells is the basis for the defective uptake of LMW
proteins in patients with
Dent's disease. The endocytosis and processing of LMW
proteins involves the multiligand tandem receptors,
megalin and
cubilin, that are abundantly expressed at the brush border of PT cells. The characterization of the endocytic defect in CLC-5 KO mice revealed that
ligands of both
megalin and
cubilin were affected, whereas a decrease in total kidney content of
megalin and
cubilin at the
protein level was detected. Using analytical subcellular fractionation and quantitative immunogold labelling, we demonstrated a selective disappearance of
megalin and
cubilin at the brush border of PT cells. These observations allowed us to conclude that defective
protein endocytosis linked to CLC-5 inactivation is due to a major and selective loss of
megalin and
cubilin at the brush border, reflecting a trafficking defect in renal PT cells. These results improve our understanding of
Dent's disease, taken as a paradigm for
renal Fanconi syndrome and
nephrolithiasis, and demonstrate multiple roles for CLC-5 in the kidney. These studies also provided insights in important functions such as apical endocytosis, handling of
proteins by renal tubular cells,
calcium metabolism, and urinary acidification.