Received for publication August 17, 2004.
Revised November 2, 2004.
Accepted for publication November 2, 2004.

Identification of amino acid residues in the insect sodium channel critical for pyrethroid binding

Abstract

The voltage-gated sodium channel is the primary target site of pyrethroids, which constitute a major class of insecticides used worldwide. Pyrethroids prolong the opening of sodium channels by inhibiting deactivation and inactivation. Despite numerous attempts to characterize pyrethroid binding to sodium channels in the past several decades, the molecular determinants of the pyrethroid-binding site on the sodium channel remain elusive. Here we show that an F to I substitution at 1519 (F1519I) in segment 6 of domain III (IIIS6) completely abolished the sensitivity of the cockroach sodium channel expressed in Xenopus oocytes to all eight structurally diverse pyrethroids examined, including permethrin and deltamethrin. In contrast, substitution by tyrosine (Y) or tryptophan (W) reduced the channel sensitivity to deltamethrin only by 3-10 fold, indicating that an aromatic residue at this position is critical for the interaction of pyrethroids with sodium channels. The F1519I mutation, however, did not alter the action of two other classes of sodium channel toxins, batrachotoxin (a site 2 toxin) and Lqh-IT (a site 3 toxin). Schild analysis utilizing competitive interaction of pyrethroid stereospecific isomers demonstrated that the F1519W mutation and a previously known pyrethroid-resistance mutation, L993F in IIS6, reduced the binding affinity of 1S cis permethrin, an inactive isomer that shares the same binding site with the active isomer 1R cis permethrin. Our results provide the first direct proof that L993 and F1519 are part of the pyrethroid receptor site on an insect sodium channel.

Key words: Sodium, Func. analysis receptor/ion channel mutants, Receptor binding studies, Receptor-mediated