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An Activation Gating Switch in Kv1.2 Is Localized to a Threonine Residue in the S2-S3 Linker

Saman Rezazadeh ^*, Harley T. Kurata ^*, Thomas W. Claydon ^*, Steven J. Kehl  and David Fedida ^*

^* Department of Anesthesiology, Pharmacology and Therapeutics, and Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada

Correspondence: Address reprint requests to Dr. David Fedida, Room 2.301, Life Sciences Centre, University of British Columbia, 2350 Health Sciences Mall, Vancouver, British Columbia V6T 1Z3, Canada. Tel.: 604-822-5806; Fax: 604-822-2316; E-mail: fedida{at}interchange.ubc.ca.

The activation properties of Kv1.2 channels are highly variable, with reported half-activation (V) values ranging from –40 mV to +30 mV. Here we show that this arises because Kv1.2 channels occupy two distinct gating modes ("fast" and "slow"). "Slow" gating (_act = 90 ± 6 ms at +35 mV) was associated with a V of activation of +16.6 ± 1.1 mV, whereas "fast" gating (_act = 4.5 ± 1.7 ms at +35 mV) was associated with a V of activation of –18.8 ± 2.3 mV. It was possible to switch between gating modes by applying a prepulse, which suggested that channels activate to a single open state along separate "fast" and "slow" activation pathways. Using chimeras and point mutants between Kv1.2 and Kv1.5 channels, we determined that introduction of a positive charge at or around threonine 252 in the S2-S3 linker of Kv1.2 abolished "slow" activation gating. Furthermore, dialysis of the cytoplasm or excision of cell-attached patches from cells expressing Kv1.2 channels switched gating from "slow" to "fast", suggesting involvement of cytoplasmic regulators. Collectively, these results demonstrate two modes of activation gating in Kv1.2 and specific residues in the S2-S3 linker that act as a switch between these modes.