title: Engineered Cx26 variants established functional heterotypic Cx26/Cx43 and Cx26/Cx40 gap junction channels.
abstract: Gap junction (GJ) channels mediate direct intercellular communication and are composed of two docked hemichannels (connexin oligomers). It is well documented that the docking and formation of GJs are possible only between compatible hemichannels (or connexins). The mechanisms of heterotypic docking compatibility are not fully clear. We aligned the protein sequences of docking-compatible and -incompatible connexins with that of connexin26 (Cx26). We found that two docking hydrogen bond (HB)-forming residues on the second extracellular domain (E2) of Cx26 and their equivalent residues are well conserved within docking-compatible connexins, but different between docking-incompatible connexins. Replacing one or both of these residues of Cx26 into the corresponding residues in the docking incompatible connexins (K168V, N176H or K168V-N176H) increased the formation of morphological and functional heterotypic GJs with connexin43 (Cx43) or connexin40 (Cx40), indicating that these two residues are important for docking incompatibility between Cx26 and these connexins. Our homology structure models predict that both HBs and hydrophobic interactions at the E2 docking interface are important docking mechanisms in heterotypic Cx26 K168V-N176H/Cx43 GJs and probably other docking compatible connexins. Revealing the key residues and mechanisms of heterotypic docking compatibility will assist us in understanding why these putative docking residues are hotspots of disease-linked mutants.
authors.affiliation: Department of Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada, N6A 5C1.
authors.forename: Levent B
authors.initials: LB
authors.lastname: Karademir
authors.affiliation: Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Aoyama
authors.affiliation: Department of Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada, N6A 5C1.
authors.forename: Benny
authors.initials: B
authors.lastname: Yue
authors.affiliation: Department of Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada, N6A 5C1.
authors.forename: Honghong
authors.initials: H
authors.lastname: Chen
authors.affiliation: Department of Physiology and Pharmacology, University of Western Ontario, London, Ontario, Canada, N6A 5C1 donglin.bai@schulich.uwo.ca.
authors.forename: Donglin
authors.initials: D
authors.lastname: Bai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D000072259:Connexin 26; D018031:Connexin 43 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017630:Connexins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004594:Electrophysiology; D017629:Gap Junctions / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D015202:Protein Engineering; D016415:Sequence Alignment; D000097003:Gap Junction alpha-5 Protein
chemical_list: D000072259:Connexin 26; D018031:Connexin 43; D017630:Connexins; D000097003:Gap Junction alpha-5 Protein; C000607042:GJB2 protein, human; C000607043:Gjb2 protein, mouse
keywords: connexin26; connexin40; connexin43; docking mechanism; gap junction channel; heterotypic docking compatibility