title: Domains responsible for constitutive and Ca(2+)-dependent interactions between calmodulin and small conductance Ca(2+)-activated potassium channels.
abstract: Small conductance Ca(2+)-activated potassium channels (SK channels) are coassembled complexes of pore-forming SK alpha subunits and calmodulin. We proposed a model for channel activation in which Ca2+ binding to calmodulin induces conformational rearrangements in calmodulin and the alpha subunits that result in channel gating. We now report fluorescence measurements that indicate conformational changes in the alpha subunit after calmodulin binding and Ca2+ binding to the alpha subunit-calmodulin complex. Two-hybrid experiments showed that the Ca(2+)-independent interaction of calmodulin with the alpha subunits requires only the C-terminal domain of calmodulin and is mediated by two noncontiguous subregions; the ability of the E-F hands to bind Ca2+ is not required. Although SK alpha subunits lack a consensus calmodulin-binding motif, mutagenesis experiments identified two positively charged residues required for Ca(2+)-independent interactions with calmodulin. Electrophysiological recordings of SK2 channels in membrane patches from oocytes coexpressing mutant calmodulins revealed that channel gating is mediated by Ca2+ binding to the first and second E-F hand motifs in the N-terminal domain of calmodulin. Taken together, the results support a calmodulin- and Ca(2+)-calmodulin-dependent conformational change in the channel alpha subunits, in which different domains of calmodulin are responsible for Ca(2+)-dependent and Ca(2+)-independent interactions. In addition, calmodulin is associated with each alpha subunit and must bind at least one Ca2+ ion for channel gating. Based on these results, a state model for Ca2+ gating was developed that simulates alterations in SK channel Ca2+ sensitivity and cooperativity associated with mutations in CaM.
authors.affiliation: Vollum Institute, Veteran's Administration Medical Center, and Department of Psychiatry, Behavioral Neuroscience, and Physiology and Pharmacology, Oregon Health Sciences University, Portland Oregon, 97201, USA.
authors.forename: J E
authors.initials: JE
authors.lastname: Keen
authors.forename: R
authors.initials: R
authors.lastname: Khawaled
authors.forename: D L
authors.initials: DL
authors.lastname: Farrens
authors.forename: T
authors.initials: T
authors.lastname: Neelands
authors.forename: A
authors.initials: A
authors.lastname: Rivard
authors.forename: C T
authors.initials: CT
authors.lastname: Bond
authors.forename: A
authors.initials: A
authors.lastname: Janowsky
authors.forename: B
authors.initials: B
authors.lastname: Fakler
authors.forename: J P
authors.initials: JP
authors.lastname: Adelman
authors.forename: J
authors.initials: J
authors.lastname: Maylie
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D002147:Calmodulin / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D005260:Female; D015640:Ion Channel Gating / Q000502:physiology; D008954:Models, Biological; D009865:Oocytes; D015221:Potassium Channels / Q000235:genetics / Q000502:physiology*; D024681:Potassium Channels, Calcium-Activated; D051381:Rats; D051640:Small-Conductance Calcium-Activated Potassium Channels; D014981:Xenopus
chemical_list: D002147:Calmodulin; C496899:Kcnn2 protein, rat; D015221:Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D051640:Small-Conductance Calcium-Activated Potassium Channels; D002118:Calcium