title: Functional characterization of renal chloride channel, CLCN5, mutations associated with Dent'sJapan disease.
abstract: BACKGROUND
  The annual urinary screening of Japanese children above three years of age has identified a progressive renal tubular disorder characterized by low molecular weight proteinuria, hypercalciuria and nephrocalcinosis, and this represents a variant of Dent's disease. Hitherto, 12 mutations of the X-linked renal specific chloride channel, CLCN5, have been reported in the Dent'sJapan variant. To further identify such CLCN5 mutations and to define the structure-function relationships of this channel, we have investigated five unrelated, non-consanguinous Japanese families with this disorder.
  
  
  METHODS
  Leukocyte DNA from probands was used with CLCN5 primers for PCR amplification of the coding region, and the DNA sequences of the products determined. Functional studies were performed by expressing the mutants in Xenopus oocytes.
  
  
  RESULTS
  Five CLCN5 mutations consisting of two nonsense (R648X and R704X), two missense (S270R and L278F) and one acceptor splice site mutation (ag-->cg) in intron 4 were identified. The missense and splice site mutations represent novel abnormalities. Heterologous expression in Xenopus oocytes of wild-type and the missense mutants demonstrated that the mutations, which were translated, either abolished or markedly reduced chloride conductance.
  
  
  CONCLUSIONS
  These results expand the spectrum of CLCN5 mutations associated with this renal disorder and provide insight into possible structure-function relationships. For example, both the missense mutations are located within a short putative loop between two transmembrane domains, and our results suggest that this region may have an important functional role in the regulation of channel activity.
authors.affiliation: Department of Pediatrics, Faculty of Medicine, University of Tokyo, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Igarashi
authors.forename: W
authors.initials: W
authors.lastname: Günther
authors.forename: T
authors.initials: T
authors.lastname: Sekine
authors.forename: J
authors.initials: J
authors.lastname: Inatomi
authors.forename: H
authors.initials: H
authors.lastname: Shiraga
authors.forename: S
authors.initials: S
authors.lastname: Takahashi
authors.forename: J
authors.initials: J
authors.lastname: Suzuki
authors.forename: N
authors.initials: N
authors.lastname: Tsuru
authors.forename: T
authors.initials: T
authors.lastname: Yanagihara
authors.forename: M
authors.initials: M
authors.lastname: Shimazu
authors.forename: T J
authors.initials: TJ
authors.lastname: Jentsch
authors.forename: R V
authors.initials: RV
authors.lastname: Thakker
mesh_terms: D000293:Adolescent; D000328:Adult; D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D002118:Calcium / Q000652:urine*; D002648:Child; D018118:Chloride Channels / Q000235:genetics* / Q000378:metabolism; D004247:DNA / Q000235:genetics; D005260:Female; D006801:Humans; D007669:Kidney Calculi / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics* / Q000502:physiology*; D009865:Oocytes / Q000378:metabolism; D010375:Pedigree; D011507:Proteinuria / Q000235:genetics*; D013577:Syndrome; D014981:Xenopus
chemical_list: C097503:CLC-5 chloride channel; D018118:Chloride Channels; D004247:DNA; D002118:Calcium