title: Conserved single residue in the BK potassium channel required for activation by alcohol and intoxication in C. elegans.
abstract: Alcohol directly modulates the BK potassium channel to alter behaviors in species ranging from invertebrates to humans. In the nematode Caenorhabditis elegans, mutations that eliminate the BK channel, SLO-1, convey dramatic resistance to intoxication by ethanol. We hypothesized that certain conserved amino acids are critical for ethanol modulation, but not for basal channel function. To identify such residues, we screened C. elegans strains with different missense mutations in the SLO-1 channel. A strain with the SLO-1 missense mutation T381I in the RCK1 domain was highly resistant to intoxication. This mutation did not interfere with other BK channel-dependent behaviors, suggesting that the mutant channel retained normal in vivo function. Knock-in of wild-type versions of the worm or human BK channel rescued intoxication and other BK channel-dependent behaviors in a slo-1-null mutant background. In contrast, knock-in of the worm T381I or equivalent human T352I mutant BK channel selectively rescued BK channel-dependent behaviors while conveying resistance to intoxication. Single-channel patch-clamp recordings confirmed that the human BK channel engineered with the T352I missense mutation was insensitive to activation by ethanol, but otherwise had normal conductance, potassium selectivity, and only subtle differences in voltage dependence. Together, our behavioral and electrophysiological results demonstrate that the T352I mutation selectively disrupts ethanol modulation of the BK channel. The T352I mutation may alter a binding site for ethanol and/or interfere with ethanol-induced conformational changes that are critical for behavioral responses to ethanol.
authors.affiliation: Waggoner Center for Alcohol and Addiction Research, Institute of Neuroscience, Center for Learning and Memory, Center for Brain, Behavior, and Evolution, and Department of Neuroscience, The University of Texas at Austin, Texas 78712.
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Davis
authors.affiliation: Waggoner Center for Alcohol and Addiction Research, Institute of Neuroscience, Center for Learning and Memory, Center for Brain, Behavior, and Evolution, and Department of Neuroscience, The University of Texas at Austin, Texas 78712.
authors.forename: Luisa L
authors.initials: LL
authors.lastname: Scott
authors.affiliation: Waggoner Center for Alcohol and Addiction Research, Institute of Neuroscience, Center for Learning and Memory, Center for Brain, Behavior, and Evolution, and Department of Neuroscience, The University of Texas at Austin, Texas 78712.
authors.forename: Kevin
authors.initials: K
authors.lastname: Hu
authors.affiliation: Waggoner Center for Alcohol and Addiction Research, Institute of Neuroscience, Center for Learning and Memory, Center for Brain, Behavior, and Evolution, and Department of Neuroscience, The University of Texas at Austin, Texas 78712 jonps@austin.utexas.edu.
authors.forename: Jonathan T
authors.initials: JT
authors.lastname: Pierce-Shimomura
mesh_terms: D000448:Aldicarb / Q000494:pharmacology; D000818:Animals; D030801:Animals, Genetically Modified; D000870:Anterior Horn Cells / Q000502:physiology; D017173:Caenorhabditis elegans; D029742:Caenorhabditis elegans Proteins / Q000235:genetics*; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics; D002492:Central Nervous System Depressants / Q000494:pharmacology*; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D007136:Immunoglobulins / Q000235:genetics; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000235:genetics*; D008124:Locomotion / Q000187:drug effects* / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense / Q000235:genetics*; D017434:Protein Structure, Tertiary / Q000235:genetics
chemical_list: D029742:Caenorhabditis elegans Proteins; D015816:Cell Adhesion Molecules, Neuronal; D002492:Central Nervous System Depressants; D002800:Cholinesterase Inhibitors; C527430:IGDCC3 protein, human; D007136:Immunoglobulins; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008164:Luminescent Proteins; C494845:slo-1 protein, C elegans; D000431:Ethanol; D000448:Aldicarb
keywords: BK channel; C. elegans; behavior; ethanol