title: Mutant connexin 50 (S276F) inhibits channel and hemichannel functions inducing cataract.
abstract: This study was designed to detect the expression, detergent resistance, subcellular localization, and channel and hemichannel functions of mutant Cx50 to understand the forming mechanism for inducing congenital cataract by a novel mutation p.S276F in connexin 50 (Cx50) reported previously by us. HeLa and human lens epithelial (HLE) cells were transfected with wild-type Cx50 and mutant Cx50 (S276F). We examined the functional characteristics of mutant Cx50 (S276F) in comparison with those of wild-type Cx50 using immunoblot, confocal fluorescence microscopy, dye transfer analysis and dye uptake assay. The mutant and wild-type Cx50 were expressed in equal levels and could efficiently localize to the plasma membrane without transportation and assembly problems. Scrape loading dye transfer was significantly evident in cells transfected with wild-type Cx50 compared to those in cells transfected with mutant Cx50 and cotransfected with wild-type and mutant Cx50. The dye uptake was found to be significantly lower in cells transfected with mutant Cx50 than in cells transfected with wild- type Cx50 and cells cotransfected with wild-type and mutant Cx50. The transfected HeLa and HLE cell lines showed similar performance in all the experiments. These results indicated that the mutant Cx50 (S276F) might inhibit the function of gap junction channel in a dominant negative manner, but inhibit the hemichannel function in a recessive negative manner.
authors.affiliation: Center for Gene Diagnosis, Zhongnan Hospital of Wuhan University, Donghu Road 169#, Wuhan, Hubei 430071, People's Republic of China. yanmingming1972@126.com.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Liu
authors.forename: Chen
authors.initials: C
authors.lastname: Qiao
authors.forename: Tanwei
authors.initials: T
authors.lastname: Wei
authors.forename: Fang
authors.initials: F
authors.lastname: Zheng
authors.forename: Shuren
authors.initials: S
authors.lastname: Guo
authors.forename: Qiang
authors.initials: Q
authors.lastname: Chen
authors.forename: Ming
authors.initials: M
authors.lastname: Yan
authors.forename: Xin
authors.initials: X
authors.lastname: Zhou
mesh_terms: D002386:Cataract / Q000235:genetics*; D004396:Coloring Agents / Q000378:metabolism; D017630:Connexins / Q000235:genetics*; D003902:Detergents / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D017629:Gap Junctions / Q000187:drug effects / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D007908:Lens, Crystalline / Q000378:metabolism; D018613:Microscopy, Confocal; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D013347:Subcellular Fractions / Q000378:metabolism; D000099264:Connexin 50
chemical_list: D004396:Coloring Agents; D017630:Connexins; D003902:Detergents; D050505:Mutant Proteins; D000099264:Connexin 50