Genetic Analyses in Dent Disease and Characterization of CLCN5 Mutations in Kidney Biopsies

Abstract

Dent disease (DD), an X-linked renal tubulopathy, is mainly caused by loss-of-function mutations in CLCN5 (DD1) and OCRL genes. CLCN5 encodes the ClC-5 antiporter that in proximal tubules (PT) participates in the receptor-mediated endocytosis of low molecular weight proteins. Few studies have analyzed the PT expression of ClC-5 and of megalin and cubilin receptors in DD1 kidney biopsies. About 25% of DD cases lack mutations in either CLCN5 or OCRL genes (DD3), and no other disease genes have been discovered so far. Sanger sequencing was used for CLCN5 gene analysis in 158 unrelated males clinically suspected of having DD. The tubular expression of ClC-5, megalin, and cubilin was assessed by immunolabeling in 10 DD1 kidney biopsies. Whole exome sequencing (WES) was performed in eight DD3 patients. Twenty-three novel CLCN5 mutations were identified. ClC-5, megalin, and cubilin were significantly lower in DD1 than in control biopsies. The tubular expression of ClC-5 when detected was irrespective of the type of mutation. In four DD3 patients, WES revealed 12 potentially pathogenic variants in three novel genes (SLC17A1, SLC9A3, and PDZK1), and in three genes known to be associated with monogenic forms of renal proximal tubulopathies (SLC3A, LRP2, and CUBN). The supposed third Dent disease-causing gene was not discovered.

Genetics and phenotypic heterogeneity of Dent disease: the dark side of the moon

“…they are genetically unsolved (Lieske et al 1993). As mentioned earlier, when NGS-either of target genes or using genetic panels-was used to investigate such cases, no other disease-causing genes were identified (Zhang et al 2017b;Anglani et al 2018;Gianesello et al 2020a).…”

Making a Dent in Dent Disease

“…Several mutations in the CBS domains have been identified in DD patients. 8 The role of ATP binding to ClC-5 in DD pathology is unclear, but ClC-5 may act as metabolic sensor that tunes V-ATPase’s ability to acidify endosomes in response to the availability of ATP and ADP. ATP binding may also be important for proper folding of ClC-5 since multiple pathogenic mutations in the CBS domains cause ER retention.…”

“… 6 Currently, over 200 different pathogenic variants of CLCN5 have been described, the majority of which result in missense or frameshift mutations with many resulting in truncation of the protein. 7 , 8 Most of the identified variants have not been functionally studied and even fewer have been studied in representative kidney cell lines or tissue. While all DD patients exhibit LMW proteinuria, the presence of other symptoms and severity of all symptoms vary greatly among patients.…”

Global gene-expression analysis reveals the molecular processes underlying ClC-5 loss-of-function in novel Dent Disease 1 cellular models

“…To date, a total of 266 pathogenic variants of CLCN5 have been reported consisting of nonsense, missense, splice site, insertion and deletion mutations [1,18]. According to the latest reports, CLCN5 mutations are grouped into three classes on the basis of functional data [16,18,19]: class 1 mutations result in defective protein processing and folding, thereby inducing retention of the mutant protein in the endoplasmic reticulum (ER), where they are early degraded by quality control systems; class 2 mutations impair protein processing and stability, leading to a functionally defective protein lacking electric currents; these mutants show reduced expression in the plasma membrane, but a normal distribution in the early endosomes; and class 3 mutations generate a protein that reaches the plasma membrane and early endosomes correctly, but shows reduced or abolished currents.…”