Reconstitution of a chemical defense signaling pathway in a heterologous system.

Chemical signaling plays an important role in ecological interactions, such as communication and predator-prey dynamics. Since sessile species cannot physically escape predators, many contain compounds that deter predation; however, it is largely unknown how predators physiologically detect deterrent chemicals. Few studies have investigated ecologically relevant aversive taste responses in any predator. Our objective was to determine if a signaling pathway for detecting marine sponge-derived deterrent compounds could be reconstituted in a heterologous expression system to ultimately facilitate investigation of the molecular mechanism of such an aversive behavioral response. Zebrafish (Danio rerio) rejected artificial diets laced with sponge chemical defense compounds that were previously shown to deter a generalist marine predator, Thalassoma bifasciatum, suggesting that zebrafish can recognize deterrent compounds relevant to coral reef systems. Transcripts made from a zebrafish cDNA library were expressed in a heterologous system, Xenopus laevis oocytes, and tested for chemoreceptor activation via electrophysiology, using the cystic fibrosis transmembrane conductance regulator (CFTR) as a reporter. Oocytes expressing gene sequences from the library and CFTR exhibited a CFTR-like electrophysiological response to formoside and ectyoplasides A and B, sponge defense compounds. Therefore, the chemical defense-activated signaling pathway can be reconstituted in Xenopus oocytes. Kinetics of the responses suggested that the responses to formoside and ectyoplasides A and B were receptor-mediated and capable of using the G(alphas) signaling pathway in this system. This bioassay has the potential to lead to the identification of genes that encode receptors capable of interacting with deterrent chemicals, which would enable understanding of predator detection of chemical defenses.