Voltage-gated sodium channels (NaV) are transmembrane proteins that are crucial for action potential initiation and propagation. They consist of a pore-forming α subunit and accessory β subunits. NaV are targeted by many toxins that directly interact with the α subunit.
We have recently discovered a new class of plant-derived toxins from Australian stinging nettles, including excelsatoxin A (ExTxA) from the giant Australian stinging tree Dendrocnide excelsa. ExTxA causes severe and long-lasting pain by activating NaV channels in sensory neurons via transmembrane protein 233 (TMEM233). Using automated patch-clamp electrophysiology in NaV/TMEM233 overexpression systems, we have characterised the selectivity of ExTxA at NaV1.1-NaV1.8 and performed detailed mechanism of action studies at NaV1.7. ExTxA profoundly inhibits NaV inactivation and decreases the time constant of recovery from inactivation. Additionally, ExTxA causes a depolarising shift in voltage-dependence of fast inactivation as well as a hyperpolarising shift in voltage-dependence of activation, and an increase in current induced by slow voltage ramps.
As virtually nothing is known about TMEM233, we next sought to investigate the effect of TMEM233 on the biophysical properties of NaV1.7 in the absence of toxin. Co-expression of TMEM233 indeed modulates electrophysiological properties of NaV1.7 by shifting both voltage-dependence of fast and slow inactivation to more hyperpolarised potentials and by increasing the time constant of recovery from inactivation.
In summary, ExTxA is the first known NaV-modulating toxin that targets an interacting protein to exert a pharmacological effect. Co-expression of TMEM233 modulates gating properties of NaV1.7 in absence of the toxin, which positions TMEM233 as a novel NaV1.7-interacting protein.