title: Connexin 32 mutations from X-linked Charcot-Marie-Tooth disease patients: functional defects and dominant negative effects.
abstract: We have characterized the function of connexin (Cx) 32 gene mutations found in X-linked dominant Charcot-Marie-Tooth disease with respect to their ability to form functional gap junctions among themselves and to inactivate wild-type Cx32 by a dominant negative mechanism. We prepared four types of Cx32 mutant cDNAs and transfected them into HeLa cells, which do not show detectable levels of gap junctional intercellular communication (GJIC), nor expression of any connexins examined. Cells transfected with the wild-type Cx32 gene, but not those transfected with three different base substitution mutations (i.e. Cys 60 to Phe, Val 139 to Met, and Arg 215 to Trp), restored GJIC. Unexpectedly, in cells transfected with a nonsense mutant at codon 220, there was also restored GJIC. When we double-transfected these mutant constructs into the HeLa cells that had already been transfected with the wild-type Cx32 gene and thus were GJIC proficient, three base substitution mutants inhibited GJIC, suggesting that these three mutants can eliminate the function of wild-type Cx32 in a dominant negative manner. The nonsense mutation at codon 220 did not show such a dominant negative effect. Since both mutant and wild-type Cx32 mRNAs were detected, but only poor Cx32 protein expression at cell-cell contact areas was observed in the double transfectants, it is suggested that certain mutants form nonfunctional chimeric connexons with wild-type connexins, which are not properly inserted into the cytoplasmic membrane.
authors.affiliation: Unit of Multistage Carcinogenesis, International Agency for Research on Cancer, Lyon, France.
authors.forename: Y
authors.initials: Y
authors.lastname: Omori
authors.forename: M
authors.initials: M
authors.lastname: Mesnil
authors.forename: H
authors.initials: H
authors.lastname: Yamasaki
mesh_terms: D000595:Amino Acid Sequence; D002450:Cell Communication / Q000235:genetics / Q000502:physiology; D002607:Charcot-Marie-Tooth Disease / Q000235:genetics*; D017630:Connexins / Q000235:genetics*; D017629:Gap Junctions / Q000502:physiology; D008040:Genetic Linkage; D006367:HeLa Cells; D006801:Humans; D009154:Mutation / Q000235:genetics*; D013347:Subcellular Fractions / Q000737:chemistry; D014960:X Chromosome / Q000235:genetics*; D000097002:Gap Junction beta-1 Protein
chemical_list: D017630:Connexins; D000097002:Gap Junction beta-1 Protein