The intracellular tetramerization domain (T1) of most eukaryotic voltage-gated potassium channels (Kv channels) exists as a “hanging gondola” below the transmembrane regions that directly control activation gating via the electromechanical coupling between the S4 voltage sensor and the main S6 gate. However, much less is known about the putative contribution of the T1 domain to Kv channel gating. This possibility is mechanistically intriguing because the T1–S1 linker connects the T1 domain to the voltage-sensing domain. Previously, we demonstrated that thiol-specific reagents inhibit Kv4.1 channels by reacting in a state-dependent manner with native Zn2+ site thiolate groups in the T1–T1 interface; therefore, we concluded that the T1–T1 interface is functionally active and not protected by Zn2+ (Wang, G., M. Shahidullah, C.A. Rocha, C. Strang, P.J. Pfaffinger, and M. Covarrubias. 2005. J. Gen. Physiol. 126:55–69). Here, we co-expressed Kv4.1 channels and auxiliary subunits (KChIP-1 and DPPX-S) to investigate the state and voltage dependence of the accessibility of MTSET to the three interfacial cysteines in the T1 domain. The results showed that the average MTSET modification rate constant (kMTSET) is dramatically enhanced in the activated state relative to the resting and inactivated states (∼260- and ∼47-fold, respectively). Crucially, under three separate conditions that produce distinct activation profiles, kMTSET is steeply voltage dependent in a manner that is precisely correlated with the peak conductance–voltage relations. These observations strongly suggest that Kv4 channel gating is tightly coupled to voltage-dependent accessibility changes of native T1 cysteines in the intersubunit Zn2+ site. Furthermore, cross-linking of cysteine pairs across the T1–T1 interface induced substantial inhibition of the channel, which supports the functionally dynamic role of T1 in channel gating. Therefore, we conclude that the complex voltage-dependent gating rearrangements of eukaryotic Kv channels are not limited to the membrane-spanning core but must include the intracellular T1–T1 interface. Oxidative stress in excitable tissues may perturb this interface to modulate Kv4 channel function.
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1 April 2006
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March 13 2006
Voltage-dependent Gating Rearrangements in the Intracellular T1–T1 Interface of a K+ Channel
Guangyu Wang,
Guangyu Wang
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College of Thomas Jefferson University, Philadelphia, PA 19107
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Manuel Covarrubias
Manuel Covarrubias
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College of Thomas Jefferson University, Philadelphia, PA 19107
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Guangyu Wang
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College of Thomas Jefferson University, Philadelphia, PA 19107
Manuel Covarrubias
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College of Thomas Jefferson University, Philadelphia, PA 19107
Correspondence to Manuel Covarrubias: [email protected]
Abbreviations used in this paper: DTT, dithiothretiol; Kv, voltage-dependent potassium; MTSET, 2-trimethylammonium-ethyl-methane-thiosulfonate bromide.
Received:
October 21 2005
Accepted:
February 15 2006
Online ISSN: 1540-7748
Print ISSN: 0022-1295
The Rockefeller University Press
2006
J Gen Physiol (2006) 127 (4): 391–400.
Article history
Received:
October 21 2005
Accepted:
February 15 2006
Citation
Guangyu Wang, Manuel Covarrubias; Voltage-dependent Gating Rearrangements in the Intracellular T1–T1 Interface of a K+ Channel . J Gen Physiol 1 April 2006; 127 (4): 391–400. doi: https://doi.org/10.1085/jgp.200509442
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