title: Regulation of ClC-2 gating by intracellular ATP.
abstract: ClC-2 is a voltage-dependent chloride channel that activates slowly at voltages negative to the chloride reversal potential. Adenosine triphosphate (ATP) and other nucleotides have been shown to bind to carboxy-terminal cystathionine-ß-synthase (CBS) domains of ClC-2, but the functional consequences of binding are not sufficiently understood. We here studied the effect of nucleotides on channel gating using single-channel and whole-cell patch clamp recordings on transfected mammalian cells. ATP slowed down macroscopic activation and deactivation time courses in a dose-dependent manner. Removal of the complete carboxy-terminus abolishes the effect of ATP, suggesting that CBS domains are necessary for ATP regulation of ClC-2 gating. Single-channel recordings identified long-lasting closed states of ATP-bound channels as basis of this gating deceleration. ClC-2 channel dimers exhibit two largely independent protopores that are opened and closed individually as well as by a common gating process. A seven-state model of common gating with altered voltage dependencies of opening and closing transitions for ATP-bound states correctly describes the effects of ATP on macroscopic and microscopic ClC-2 currents. To test for a potential pathophysiological impact of ClC-2 regulation by ATP, we studied ClC-2 channels carrying naturally occurring sequence variants found in patients with idiopathic generalized epilepsy, G715E, R577Q, and R653T. All naturally occurring sequence variants accelerate common gating in the presence but not in the absence of ATP. We propose that ClC-2 uses ATP as a co-factor to slow down common gating for sufficient electrical stability of neurons under physiological conditions.
authors: Institute of Complex Systems-Zelluläre Biophysik (ICS-4), Forschungszentrum Jülich, 52425, Jülich, Germany, g.stoelting@fz-juelich.de.
authors: Gabriel
authors: G
authors: Stölting
authors: Georgeta
authors: G
authors: Teodorescu
authors: Birgit
authors: B
authors: Begemann
authors: Julian
authors: J
authors: Schubert
authors: Rima
authors: R
authors: Nabbout
authors: Mohammad Reza
authors: MR
authors: Toliat
authors: Thomas
authors: T
authors: Sander
authors: Peter
authors: P
authors: Nürnberg
authors: Holger
authors: H
authors: Lerche
authors: Christoph
authors: C
authors: Fahlke
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000090007:CLC-2 Chloride Channels; D018118:Chloride Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004829:Epilepsy, Generalized / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism; D015640:Ion Channel Gating; D020125:Mutation, Missense; D017434:Protein Structure, Tertiary
chemical_list: D000090007:CLC-2 Chloride Channels; D018118:Chloride Channels; D000255:Adenosine Triphosphate
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