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 (PIP ₂ ) 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.