Pore stoichiometry of a voltage-gated chloride channel.
Ion channels allow ions to pass through cell membranes by forming aqueous permeation pathways (pores). In contrast to most known ion channels, which have single pores, a chloride channel belonging to the CIC family (Torpedo CIC-0) has functional features that suggest that it has a unique 'double-barrelled' architecture in which each of two subunits forms an independent pore. This model is based on single-channel recordings of CIC-0 that has two equally spaced and independently gated conductance states. Other CIC isoforms do not behave in this way, raising doubts about the applicability of the model to all CIC channels. Here we determine the pore stoichiometry of another CIC isoform, human CIC-1, by chemically modifying cysteines that have been substituted for other amino acids located within the CIC ion-selectivity filter. The CIC-1 channel can be rendered completely susceptible to block by methanethiosulphonate reagents when only one of the two subunits contains substituted cysteines. Thiol side chains placed at corresponding positions in both subunits can form intersubunit disulphide bridges and coordinate Cd2+, indicating that the pore-forming regions from each subunit line the same conduction pathway. We conclude that human CIC-1 has a single functional pore.
Department of Medicine (Nephrology), and The Center for Molecular Neuroscience, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-6600, USA. christoph.fahlke@mcmail.vanderbilt.edu
C
C
Fahlke
T H
TH
Rhodes
R R
RR
Desai
A L
AL
George
D002104:Cadmium / Q000494:pharmacology; D002460:Cell Line; D018118:Chloride Channels / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D004563:Electrochemistry; D006639:Histidine / Q000737:chemistry; D006801:Humans; D015640:Ion Channel Gating; D008698:Mesylates / Q000494:pharmacology; D008968:Molecular Conformation; D009124:Muscle Proteins / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010618:Phenanthrolines / Q000494:pharmacology; D013438:Sulfhydryl Compounds / Q000737:chemistry
C086321:CLC-1 channel; D018118:Chloride Channels; D008698:Mesylates; D009124:Muscle Proteins; D010618:Phenanthrolines; D013438:Sulfhydryl Compounds; D002104:Cadmium; C109153:(2-sulfonatoethyl)methanethiosulfonate; C109154:(2-(trimethylammonium)ethyl)methanethiosulfonate; C014741:bis(1,10-phenanthroline)copper(2+) ion; D006639:Histidine; D003545:Cysteine