Seiferth, D. | Search bioRxiv

bioRxiv2025

Structural determinants for GqPCR-mediated inhibition of TASK K2P K+ channels and their dysfunction in disease

Two-Pore Domain K+(K2P) channels are crucial determinants of cellular electrical excitability. TASK-1 and TASK-3 K2P channels regulate the resting membrane potential in many different cell types where their activity is also coupled to GqPCR signalling pathways via direct inhibition by diacylglycerol (DAG) generated as a result of phosphatidylinositol-4,5-bisphosphate (PIP2) hydrolysis. This regulation is defective in two different TASK channelopathies, but the molecular mechanisms underlying this inhibition remain unclear. Here, we demonstrate that DAG inhibition of TASK channel activity is state-dependent. Single channel recordings show that the sensitivity to GqPCR inhibition inversely correlates with channel open probability and that DAG destabilises the open state of TASK-1 to promote channel closure. Combining Molecular Dynamics simulations with mutagenesis studies, we also identify a binding site for DAG in a groove between the M2, M3 and M4 domains, and highlight the crucial role of a specific residue within on M4 (T230) in mediating this inhibitory effect as well as defining the difference in GPCR sensitivity between TASK-1 and TASK-3. Together, these results provide a better understanding of the molecular mechanisms underlying GqPCR regulation of TASK channels and the pathogenic effect of K2P channelopathies linked to TASK channel dysregulation.

biophysics↗

bioRxiv2024

Exploring the Influence of Pore Shape on Conductance and Permeation

There are increasing numbers of ion channel structures featuring heteromeric subunit assembly, exemplified by synaptic 1{beta}B Glycine and 4{beta}2 Nicotinic receptors. These structures exhibit inherent pore asymmetry, but the relevance of this to function is unknown. Furthermore, molecular dynamics simulations performed on symmetrical homomeric channels often leads to thermal distortion whereby conformations of the resulting ensemble are also asymmetrical. When functionally annotating ion channels, researchers often rely on minimal constrictions determined via radius-profile calculations performed with computer programs, such as HOLE or CHAP, coupled with an assessment of pore hydrophobicity. However, such tools typically employ spherical probe particles, limiting their ability to accurately capture pore asymmetry. Here, we introduce an algorithm that employs ellipsoidal probe particles, enabling a more comprehensive representation of the pore geometry. Our analysis reveals that the use of non-spherical ellipsoids for pore characterization, provides a more accurate and easily interpretable depiction of conductance. To quantify the implications of pore asymmetry on conductance, we systematically investigated carbon nanotubes (CNTs) with varying degrees of pore asymmetry as model systems. The conductance through these channels shows surprising effects that would otherwise not be predicted with spherical probes. The results have broad implications not only for the functional annotation of biological ion channels, but also for the design of synthetic channel systems for use in areas such as water filtration. Furthermore, we make use of the more accurate characterization of channel pores to refine a physical conductance model to obtain a heuristic estimate for single channel conductance. The code is freely available, obtainable as pip-installable python package and provided as a webservice.

biophysics↗