Professor Stephen Tucker

Abstract:

Two-Pore Domain K+ (K2P) channels are crucial determinants of the resting membrane potential and of cellular electrical excitability in many different cell types. TASK-1 and TASK-3 K2P channel activity is also coupled to GPCR signalling pathways via Gαq and their subsequent inhibition is via direct interaction with diacylglycerol (DAG) generated from phosphatidylinositol-4,5-bisphosphate (PIP₂) hydrolysis. This regulation is defective in two different neurodevelopmental disorders, but the molecular mechanisms underlying this inhibitory process and the reasons for the GPCR-insensitivity of these disease-causing mutations remain unclear. Here we show that GqPCR inhibition inversely correlates with channel open probability, and results from a state-dependent destabilisation of the open state by DAG promoting channel closure. We also identify a DAG interaction-site within a groove between the M2, M3 and M4 domains, and show the crucial role of residues within this site in mediating the inhibitory effect and defining channel sensitivity. These results not only reveal the structural and molecular mechanisms underlying GqPCR regulation of TASK channels, but also explain the pathogenic effect of a common regulatory defect linked to different K2P channelopathies.

Abstract:

Pain management in animal experimentation is crucial for both ethical and scientific reasons, as unmanaged pain can distort physiological responses compromising data reliability. Current strategies are often invasive and pharmacology-based, introducing variability and confounding effects. Here, we present Light-Induced Analgesia, a drug-free, non-invasive method for pain relief in animals. We show that 365 nm illumination activates the pain-inhibitory TRAAK two-pore domain potassium (K2P) channel. This activation is driven by the oxidation of a native methionine at TRAAK’s regulatory fenestration site, triggering a conformational switch from its inactive (down) to active (up) state. We further demonstrate that this mechanism can be transferred to other related K2Ps via a single-point mutation, rendering them light-sensitive. In rodents, gentle skin exposure to 365 nm is sufficient to activate endogenous TRAAK, silence nociceptors, and produce potent, long-lasting analgesia that outperforms standard treatments. Light-Induced Analgesia thus offers an effective, drug-free alternative that can enhance animal welfare and experimental reliability in preclinical research.

Abstract:

The 2TM domain family of K channels are also known as the inward-rectifier K channel family. This family includes the strong inward-rectifier K channels (Kir2.x) that are constitutively active, the G protein-activated inward-rectifier K channels (Kir3.x) and the ATP-sensitive K channels (Kir6.x, which combine with sulphonylurea receptors (SUR1-3)). The pore-forming α subunits form tetramers, and heteromeric channels may be formed within subfamilies (e.g. Kir3.2 with Kir3.3).

Abstract:

The 2TM domain family of K channels are also known as the inward-rectifier K channel family. This family includes the strong inward-rectifier K channels (Kir2.x) that are constitutively active, the G-protein-activated inward-rectifier K channels (Kir3.x) and the ATP-sensitive K channels (Kir6.x, which combine with sulphonylurea receptors (SUR1-3)). The pore-forming α subunits form tetramers, and heteromeric channels may be formed within subfamilies (e.g. Kir3.2 with Kir3.3).

Abstract:

THIK-1 (KCNK13) is a halothane-inhibited and anionic-lipid-activated two-pore domain (K2P) K+ channel implicated in microglial activation and neuroinflammation, and a current target for the treatment of neurodegenerative disorders, for example Alzheimer’s disease and amyothropic lateral sclerosis (ALS). However, compared to other K2P channels, little is known about the structural and functional properties of THIK-1. Here we present a 3.16-Å-resolution cryo-EM structure of human THIK-1 that reveals several distinct features, in particular, a tyrosine in M4 that contributes to a lower ‘Y gate’ that opens upon activation by physiologically relevant G-protein-coupled receptor and lipid signaling pathways. We demonstrate that linoleic acid bound within a modulatory pocket adjacent to the filter influences channel activity, and that halothane inhibition involves a binding site within the inner cavity, both resulting in conformational changes to the Y gate. Finally, the extracellular cap domain contains positively charged residues that line the ion exit pathway and contribute to the distinct biophysical properties of this channel. Overall, our results provide structural insights into THIK-1 function and identify distinct regulatory sites that expand its potential as a drug target for the modulation of microglial function.