BPS2026 – The mechanism of Palythoa’s resistance to palytoxin

Palytoxin (PTX) is a sodium pump-specific toxin originally extracted from cnidarian zoanthids of the genus Palythoa . PTX binds at the extracellular surface of sodium pumps from every known animal source, opening a cation channel through the pump molecule. All animals, from sponges to vertebrates, express at least one form of the sodium pump, which is composed of a catalytic α- and an auxiliary β-subunit. Do Palythoa have PTX-resistant sodium pumps? We used a 2-electrode voltage clamp (TEVC) after expression in Xenopus oocytes to test if pumps from the cnidarian Hydra vulgaris presented normal function and were sensitive to PTX. K-activated outward currents in oocytes injected with Hydra pumps and 10 pM PTX application in Na solution activated large inward current at −50 mV. We identified sodium pump α and β-subunits within Palythoa transcriptomes. Palythoa α-subunit co-injected with Palythoa , human, or Hydra β-subunits failed to form functional pumps. We constructed chimeras in which the transmembrane regions (M1⋯M10) and extracellular loops of the Palythoa caribaeorum α-subunit were substituted within the corresponding regions of the human α1. These α-subunit chimeras were expressed with human β1 and their function and PTX sensitivity were studied with TEVC and patch clamp. Two sections of the catalytic subunit were found to reduce the apparent affinity for palytoxin: The M3–M4 chimera reduced affinity by ∼50-fold, from ∼50 pM to ∼2 nM, while M7–M8 reduced it by ∼ 10-fold. A chimera combining these two regions produced pumps with normal function, that were insensitive to up to 10 nM PTX, consistent with a ∼500-fold reduction in PTX affinity for the sodium pump. These results indicate that these regions are essential for PTX binding and action on the sodium pump and suggest that Palythoa survive high PTX concentrations by expressing PTX-insensitive sodium pumps.