A novel mutation from gene splicing of a voltage-gated sodium channel in a marine copepod and its potential effect on channel function

The saxitoxins (STX), a group of potent neurotoxins produced by some marine algae (dinoflagellates and cyanobacteria), block voltage-gated sodium channels, inhibiting nerve-signal transmission in consumers of STX-bearing prey. Populations of grazers (clams and copepods) persistently exposed to the STX-bearing dinoflagellate Alexandrium fundyense are less susceptible to STX than naive ones. Adaptation to STX in clams is linked to a point mutation at the STX-binding site in the sodium channel, which dramatically lowers the sensitivity to the toxin (STX resistance). The present study tested if a similar mechanism of STX resistance occurs in the copepod Acartia hudsonica. Our cloning and sequencing results indicate that two full-length cDNA variants (AhNa(v)1 and AhNa(v)2) of the sodium channel exist in A. hudsonica, which result from alternative splicing of the single coding gene. Both variants have identical nucleotide sequences except that AhNa(v)1 (the putative mutant isoform) contains a three-amino-acid (GRD) insertion and a single adjacent aa-substitution (A to V) close to the inactivation gate on the cytoplasmic linker between domains III and IV of the sodium channel. All individuals express both AhNa(v)1 and AhNa(v)2 in varying proportions. The functional consequences of the mutation were studied by inserting the three-amino acid codons into a rat (rNa(v)31.2) sodium channel expressed in both Xenopus oocytes and HEK cells. Currents carried by construct rNa(v)1.2 bearing the GRD insertion did not inactivate as completely, and recovered faster from inactivation than rNa(v)1.2. These two rNa(v)1.2 constructs were, however, equally sensitive to STX, suggesting that the GRD variation does not confer STX resistance on the rat sequence of Na(v)1.2. These results render unlikely the hypothesis that this novel mutation is responsible for the adaptation (via resistance) of A. hudsonica to STX-bearing prey. (C) 2015 Elsevier B.V. All rights reserved.