title: A missense glial cells missing homolog B (GCMB) mutation, Asn502His, causes autosomal dominant hypoparathyroidism.
abstract: BACKGROUND
  Glial cells missing B (GCMB), the mammalian homolog of the Drosophila GCM gene, encodes a 506-amino-acid parathyroid-specific transcription factor. To date, only two different heterozygous GCMB mutations have been reported in three kindreds with autosomal dominant hypoparathyroidism.
  
  
  OBJECTIVE
  Our objective was to investigate a family with autosomal dominant hypoparathyroidism for PTH, CaSR, and GCMB mutations.
  
  
  METHODS
  Leukocyte DNA was used with exon-specific primers for PCR amplification and the DNA sequences of the PCR products determined. Functional analyses using fluorescence microscopy, EMSAs, and luciferase reporter assays were undertaken. Informed consent was obtained using protocols approved by a national ethical committee.
  
  
  RESULTS
  DNA sequence analysis revealed an A to C transversion at codon 502 of GCMB, which altered the wild-type asparagine (Asn) to histidine (His). Functional studies, using transient transfections of COS7 cells with GCMB wild-type and mutant (Asn502His) tagged constructs, demonstrated that the wild-type and mutant proteins localized to the nucleus and retained the ability to bind the GCM-consensus DNA recognition motif. However, a luciferase reporter assay demonstrated that the Asn502His mutation resulted in a reduction in gene transactivation. Moreover, cotransfection of the wild-type with mutant did not lead to an increase in luciferase activity, thereby demonstrating a dominant-negative effect of the Asn502His mutant that would be consistent with an autosomal dominant inheritance.
  
  
  CONCLUSIONS
  Our results, which have identified the first dominant missense GCMB mutation, help to increase our understanding of the mechanism underlying gene transactivation that is a prerequisite for the function of this parathyroid gland-specific transcription factor.
authors.affiliation: Academic Endocrine Unit, Nuffield Department of Clinical Medicine, University of Oxford, Oxford Centre for Diabetes, Endocrinology, and Metabolism (OCDEM), Churchill Hospital, Headington Oxford OX3 7LJ, UK.
authors.forename: Samantha M
authors.initials: SM
authors.lastname: Mirczuk
authors.forename: Michael R
authors.initials: MR
authors.lastname: Bowl
authors.forename: M Andrew
authors.initials: MA
authors.lastname: Nesbit
authors.forename: Treena
authors.initials: T
authors.lastname: Cranston
authors.forename: Carl
authors.initials: C
authors.lastname: Fratter
authors.forename: Jeremy
authors.initials: J
authors.lastname: Allgrove
authors.forename: Caroline
authors.initials: C
authors.lastname: Brain
authors.forename: Rajesh V
authors.initials: RV
authors.lastname: Thakker
mesh_terms: D015153:Blotting, Western; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007011:Hypoparathyroidism / Q000235:genetics*; D008856:Microscopy, Fluorescence; D020125:Mutation, Missense / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics*
chemical_list: C119532:GCM2 protein, human; D009687:Nuclear Proteins; D014157:Transcription Factors