title: Different mechanisms preclude mutant CLDN14 proteins from forming tight junctions in vitro.
abstract: Mutations in claudin 14 (CLDN14) cause nonsyndromic DFNB29 deafness in humans. The analysis of a murine model indicated that this phenotype is associated with degeneration of hair cells, possibly due to cation overload. However, the mechanism linking these alterations to CLDN14 mutations is unknown. To investigate this mechanism, we compared the ability of wild-type and missense mutant CLDN14 to form tight junctions. Ectopic expression in L mouse fibroblasts (LM cells) of wild-type CLDN14 protein induced the formation of tight junctions, while both the c.254T>A (p.V85D) mutant, previously identified in a Pakistani family, and the c.301 G>A (p.G101R) mutant, identified in this study through the screen of 183 Spanish and Greek patients affected with sporadic nonsyndromic deafness, failed to form such junctions. However, the two mutant proteins differed in their ability to localize at the plasma membrane. We further identified hitherto undescribed exons of CLDN14 that are utilized in alternative spliced transcripts. We demonstrated that different mutations of CLDN14 impaired by different mechanisms the ability of the protein to form tight junctions. Our results indicate that the ability of CLDN14 to be recruited to these junctions is crucial for the hearing process.
authors.affiliation: Department of Genetic Medicine and Development, University of Geneva Medical School and Geneva University Hospitals, Geneva, Switzerland.
authors.forename: Marie
authors.initials: M
authors.lastname: Wattenhofer
authors.forename: Alexandre
authors.initials: A
authors.lastname: Reymond
authors.forename: Véronique
authors.initials: V
authors.lastname: Falciola
authors.forename: Anne
authors.initials: A
authors.lastname: Charollais
authors.forename: Dorothée
authors.initials: D
authors.lastname: Caille
authors.forename: Christelle
authors.initials: C
authors.lastname: Borel
authors.forename: Robert
authors.initials: R
authors.lastname: Lyle
authors.forename: Xavier
authors.initials: X
authors.lastname: Estivill
authors.forename: Michael B
authors.initials: MB
authors.lastname: Petersen
authors.forename: Paolo
authors.initials: P
authors.lastname: Meda
authors.forename: Stylianos E
authors.initials: SE
authors.lastname: Antonarakis
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D057167:Claudins; D004285:Dogs; D005091:Exons / Q000235:genetics; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D007438:Introns / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D019108:Tight Junctions / Q000737:chemistry / Q000378:metabolism*
chemical_list: D057167:Claudins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C422064:claudin 14