title: Intracellular distribution, assembly and effect of disease-associated connexin 31 mutants in HeLa cells.
abstract: Mutations in connexin 31 (Cx31) are associated with erythrokeratodermia variabilis (EKV), hearing impairment and peripheral neuropathy; however, the pathological mechanism of Cx31 mutants remains unknown. This study analyzed 11 disease-associated Cx31 variants and one non-disease-associated Cx31 variant and compared their intracellular distribution and assembly in HeLa cells and their effect on these cells. The fluorescent localization assay showed no gap junction plaque formation in the cells expressing the recessive EKV-associated mutant (L34P) and four hearing impairment-associated mutants (66delD, 141delI, R180X and E183K), significantly reduced plaque formation in the cells with five EKV-associated dominant mutants (G12R, G12D, R42P, C86S and F137L) and no obvious change in the cells with two other mutants (I141V and 652del12). Immunoblotting analysis showed that 12 mutated Cx31s, like WT-Cx31, are able to form the Triton X-100 insoluble complex; however, the quantity of Triton X-100 insoluble complex in the transfected HeLa cells varied among different Cx31 mutants. Additionally, the expression of five EKV-associated dominant mutants (G12R, G12D, R42P, C86S and F137L) caused cell death in HeLa cells. However, the five hearing impairment-associated mutants did not induce cell death. The above results suggest that disease-associated mutants gain deleterious functions differentially. In summary, disease-associated Cx31 mutants impair the formation of normal gap junctions at different levels, and the diseases associated with Cx31 mutations may result from the abnormal assembly, trafficking and metabolism of the Cx31 mutants.
authors.affiliation: National Laboratory of Medical Genetics, Central South University, Changsha 410078, China.
authors.forename: Li-Qiang
authors.initials: LQ
authors.lastname: He
authors.forename: Yu
authors.initials: Y
authors.lastname: Liu
authors.forename: Fang
authors.initials: F
authors.lastname: Cai
authors.forename: Zhi-Ping
authors.initials: ZP
authors.lastname: Tan
authors.forename: Qian
authors.initials: Q
authors.lastname: Pan
authors.forename: De-Sheng
authors.initials: DS
authors.lastname: Liang
authors.forename: Zhi-Gao
authors.initials: ZG
authors.lastname: Long
authors.forename: Ling-Qian
authors.initials: LQ
authors.lastname: Wu
authors.forename: Liang-Qun
authors.initials: LQ
authors.lastname: Huang
authors.forename: He-Ping
authors.initials: HP
authors.lastname: Dai
authors.forename: Kun
authors.initials: K
authors.lastname: Xia
authors.forename: Jia-Hui
authors.initials: JH
authors.lastname: Xia
authors.forename: Zhuo-Hua
authors.initials: ZH
authors.lastname: Zhang
mesh_terms: D017209:Apoptosis; D017630:Connexins / Q000235:genetics* / Q000378:metabolism*; D004890:Erythema / Q000235:genetics / Q000378:metabolism*; D017629:Gap Junctions / Q000378:metabolism*; D006367:HeLa Cells; D006311:Hearing Disorders / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007642:Keratosis / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010523:Peripheral Nervous System Diseases / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D011994:Recombinant Proteins / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D017630:Connexins; D011994:Recombinant Proteins; C068044:GJB3 protein, human