title: Tandem pore domain halothane-inhibited K+ channel subunits THIK1 and THIK2 assemble and form active channels.
abstract: Despite a high level of sequence homology, tandem pore domain halothane-inhibited K(+) channel 1 (THIK1) produces background K(+) currents, whereas THIK2 is silent. This lack of activity is due to a unique combination of intracellular retention and weak basal activity in the plasma membrane. Here, we designed THIK subunits containing dominant negative mutations (THIK1(DN) and THIK2(DN)). THIK2(DN) mutant inhibits THIK1 currents, whereas THIK1(DN) inhibits an activated form of THIK2 (THIK2-A155P-I158D). In situ proximity ligation assays and Förster/fluorescence resonance energy transfer (FRET) experiments support a physical association between THIK1 and THIK2. Next, we expressed covalent tandems of THIK proteins to obtain expression of pure heterodimers. Td-THIK1-THIK2 (where Td indicates tandem) produces K(+) currents of amplitude similar to Td-THIK1-THIK1 but with a noticeable difference in the current kinetics. Unlike Td-THIK2-THIK2 that is mainly detected in the endoplasmic reticulum, Td-THIK1-THIK2 distributes at the plasma membrane, indicating that THIK1 can mask the endoplasmic reticulum retention/retrieval motif of THIK2. Kinetics and unitary conductance of Td-THIK1-THIK2 are intermediate between THIK1 and THIK2. Altogether, these results show that THIK1 and THIK2 form active heteromeric channels, further expanding the known repertoire of K(+) channels.
authors.affiliation: From LabEx ICST, Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France and.
authors.forename: Sandy
authors.initials: S
authors.lastname: Blin
authors.affiliation: From LabEx ICST, Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France and.
authors.forename: Franck C
authors.initials: FC
authors.lastname: Chatelain
authors.affiliation: From LabEx ICST, Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France and.
authors.forename: Sylvain
authors.initials: S
authors.lastname: Feliciangeli
authors.affiliation: the Department of Physiology and Institute of Health Sciences, Gyeongsang National University, School of Medicine, Jinju 660-751, South Korea.
authors.forename: Dawon
authors.initials: D
authors.lastname: Kang
authors.affiliation: From LabEx ICST, Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France and lesage@ipmc.cnrs.fr.
authors.forename: Florian
authors.initials: F
authors.lastname: Lesage
authors.affiliation: From LabEx ICST, Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, and Université de Nice Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France and.
authors.forename: Delphine
authors.initials: D
authors.lastname: Bichet
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002712:Chlorides / Q000737:chemistry* / Q000378:metabolism; D004285:Dogs; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D017136:Ion Transport; D007700:Kinetics; D061985:Madin Darby Canine Kidney Cells; D008564:Membrane Potentials; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009865:Oocytes / Q000166:cytology / Q000502:physiology; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000737:chemistry* / Q000378:metabolism; D024683:Potassium Channels, Tandem Pore Domain / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
chemical_list: D002712:Chlorides; C586367:KCNK12 protein, human; C486603:KCNK13 protein, human; D024683:Potassium Channels, Tandem Pore Domain; D011994:Recombinant Proteins; D011188:Potassium
keywords: Electrophysiology; K2P Channels; KCNK; Plasma Membrane; Potassium Channel; Protein Assembly; Protein Complex