Molecular analysis of the patients with idiopathic low molecular weight proteinuria

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Chloride channel 5 protein, CLC5, is highly expressed in the cytosomes and lysosomal membrane of renal proximal tubules which works to keep the intralysosomal pH low. We have indicated the clinical similarity between Japanese idiopathic low molecular weight proteinuria and British Dent's disease, and identified 18 different mutations of CLC5 gene (CLCN5) in 21 families among 29 families with idiopathic low molecular weight proteinuria. Three nonsense mutations, W270X, R648X and R704X, found in 5 Japanese families of the disease was also found in British families with Dent's disease and North American X-linked nephrolithiasis suggesting that those mutations are mutational hot spots in CLCN5. Seven mutations (33%) occurred in exon 8 of CLCN5, and 4 (19%) in exon 11. Heterologous expression of 4 missense mutations, S270R, L278F, R280P and L706P in Xenopus oocytes demonstrated 70%, 100%, 70% and 30% reduction respectively in channel activity when compared with the wild-type. Western blot analysis disclosed that the mutated protein L706P is expressed not only in oocyte membrane but in cytoplasm suggesting that the terminal portion of CLC5 protein might act as an anchoring the protein at the cell membrane. The genotype phenotype correlation analyzed by Man-Whitney test revealed that 1) urine beta2-microglobulin and Ca/Cr are elevated in the patients with CLCN5 mutations than in the controls, 2) urine Ca/Cr is lower in the patients with CLCN5 missense mutations than in the patients with other CLCN5 mutations, and 3) the incidence of nephrocalcinosis is lower in patients with CLCN5 missense mutations than in the patients with other CLCN5 mutations.