Allosteric coupling between proximal C-terminus and selectivity filter is facilitated by the movement of transmembrane segment 4 in TREK-2 channel

Abstract

TREK-2, a member of two-pore-domain potassium channel family, regulates cellular excitability in response to diverse stimuli. However, how such stimuli control channel function remains unclear. Here, by characterizing the responses of cytosolic proximal C-terminus deletant (ΔpCt) and transmembrane segment 4 (M4)-glycine hinge mutant (G312A) to 2-Aminoethoxydiphenyl borate (2-APB), an activator of TREK-2, we show that the transduction initiated from pCt domain is allosterically coupled with the conformation of selectivity filter (SF) via the movements of M4, without depending on the original status of SF. Moreover, ΔpCt and G312A also exhibited blunted responses to extracellular alkalization, a model to induce SF conformational transition. These results suggest that the coupling between pCt domain and SF is bidirectional, and M4 movements are involved in both processes. Further mechanistic exploration reveals that the function of Phe316, a residue close to the C-terminus of M4, is associated with such communications. However, unlike TREK-2, M4-hinge of TREK-1 only controls the transmission from pCt to SF, rather than SF conformational changes triggered by pHo changes. Together, our findings uncover the unique gating properties of TREK-2, and elucidate the mechanisms for how the extracellular and intracellular stimuli harness the pore gating allosterically.

Gating, Regulation, and Structure in K2P K+ Channels: In Varietate Concordia?

“…Perhaps surprisingly, 2-APB effects were not dependent on extracellular K 1 concentration. Zhuo et al (2016) interpreted their results in terms of an allosteric coupling of stimuli sensed at intracellular portions of the channel structure and gating at the SF, which supports the hypothesis of SF activation or inactivation as the sole mode of K 2P channel gating.…”

“…Recent work by Zhuo et al (2016) has addressed the gating mechanism for activation of TREK-2 by 2-aminoethoxydiphenyl borate (2-APB). The 2-APB site of action is a proximal portion of the C terminus (pCt) that, when deleted, as in mutant DpCt, leads to a loss of 2-APB-induced activity.…”

Antagonistic Effect of a Cytoplasmic Domain on the Basal Activity of Polymodal Potassium Channels

“…An allosteric coupling between pCt and the selectivity filter (SF) has been described in TREK2. A movement of M4 facilitates this coupling ( Zhuo et al, 2016 ). To investigate the possibility that coupling is necessary for pCt action in TREK1 and TRAAK, we modified the M4/pCt junction by replacing residues 292 to 294 by glycines in TREK1 to produce TREK1-3G, and residues 253 to 255 in TRAAK to produce TRAAK-3G.…”

Intersubunit Concerted Cooperative and cis-Type Mechanisms Modulate Allosteric Gating in Two-Pore-Domain Potassium Channel TREK-2

“…Our previous studies have demonstrated that 2-Aminoethoxydiphenyl borate (2-APB), a membrane-permeable compound, stimulates the activity of TREK-2 via acting on its cytosolic proximal C-terminus (pCt; Zhuo et al, 2015a ), and opens the pore regardless of the original conformation of SF gate (Zhuo et al, 2016 ). This allosteric transduction (named as “2-APB pathway”) is initiated from cytosolic Ct and propagated to SF gate.…”

“…In addition, changes of extracelluar pH (ΔpH o ) induce the conformational transition of SF gate in K2Ps via an external sensor (Clarke et al, 2008 ; Cohen et al, 2008 ; Stansfeld et al, 2008 ; Niemeyer et al, 2010 ; Ma et al, 2011 ; Zúñiga et al, 2011 ), including TREK-2 (Sandoz et al, 2009 ; Zhuo et al, 2015a ). Such transition in TREK-2 is also facilitated by the pCt domain in TREK-2 (Zhuo et al, 2016 ). This transmission is evoked by the charge change of sensor and propagated such stimuli to SF gate, and ultimately to intracellular pCt domain, a process called as “pH o pathway”.…”