Barro Soria, R. | bioRxiv Science

bioRxiv2025

Structural basis of ATP release and ion selectivity in Pannexin1 channels

This study investigated the structural basis of adenosine triphosphate (ATP) release and ion selectivity by Pannexin1 (Panx1). The microinjection of active SRC tyrosine kinase increased Panx1-mediated currents in Panx1-expressing oocytes. Furthermore, a mutation mimicking phosphorylation at Y308 turned Panx1 into a constitutively open ATP-releasing channel with broad ion permeability. Cryogenic electron microscopy showed that for most maps, the channel alternated between wide and narrow conformations. However, in the map of the phosphomimetic Y308E mutation, Panx1 remains in a wide ATP-releasing conformation. The pore-lining residues, W74 and R75, are flexible and create, in concert with the sequestered N-terminal helix, a path for ATP. The data point to phosphorylation as a reversible switch that activates ATP permeability by stabilizing Panx1 in the ATP-releasing conformation, which is crucial for developing modulators for diseases such as cancer and chronic pain.

biophysics↗

bioRxiv2022

Distinctive mechanisms of epilepsy-causing mutants discovered by measuring S4 movement in KCNQ2 channels

Neuronal KCNQ channels mediate the muscarine-regulated M-current, a key regulator of membrane excitability in the central and peripheral nervous systems. Mutations in KCNQ2 channels cause severe neurodevelopmental disorders, including epileptic encephalopathies. However, the impact that mutations have on channel function remain poorly defined, largely because of our limited understanding of the voltage sensing mechanisms that trigger channel gating. Here, we present measurements of voltage sensor movements in wt-KCNQ2 and channels bearing epilepsy-causing mutations using mutagenesis, cysteine accessibility, and voltage clamp fluorometry (VCF). Cysteine modification reveals that a stretch of 8-9 amino acids in the S4 become exposed upon opening of KCNQ2 channels. VCF shows that the voltage dependence and kinetics of S4 movement and channel opening/closing closely correlate, suggesting an activation scheme in which channel opening does not require multiple voltage-sensor movements. VCF and kinetic modeling reveal different mechanisms by which epilepsy-causing mutations affect KCNQ2 channel voltage-dependent gating. This study provides insight into KCNQ2 channel function, which will aid in uncovering the mechanisms underlying channelopathies.

biophysics↗