Sequence, characterization and pharmacological analyses of the adipokinetic hormone receptor in the stick insect,

Background: Adipokinetic/hypertrehalosaemic hormone (AKH/HrTH), corazonin (Crz) and the AKH/Crz-related peptide (ACP) are neuropeptides considered homologous to the vertebrate gonadotropin-releasing hormone (GnRH). AKH/HrTH are important peptidergic metabolic regulators in insects that are crucial to provide energy during periods of high output mobility or when large amounts of energy-rich substrates are synthesized (for example, during vitellogenesis). AKH functions via a G protein-coupled receptor. Understanding which residue of the peptide (the ligand), activates the receptor with high efficacy is an important step to get insights into the ligand-receptor interaction, which is essential for further research on creating a model of how the ligand behaves in the binding pocket of the receptor. Such data are necessary for the search of non-peptidic mimetic agonists or antagonists in pesticide design. Methods: Using bioinformatics and cloning techniques, the complete coding sequence of an AKH receptor was cloned and sequenced from fat body tissues and nervous tissues from the Indian stick insect, Results: The stick insect AKH receptor was successfully cloned and sequenced from fat body and, separately, from nervous tissues. Comparison with known insect AKH, Crz and ACP receptors clearly put the stick insect receptor in the AKH clade and as sister group to other putative Phasmatodean AKH receptors. Moreover, the receptor expressed in mammalian cells was only activated by AKH and not by Crz or ACP indicating a true AKH receptor. Structure-activity studies in an Ala replacement series revealed the ligand residues that are absolutely essential for activating the AKHR: the N-terminal pGlu, Phe Conclusions: The AKH receptor from stick insects (Phasmatodea) cluster together in one clade distinct from other insect AKHRs, although still similar enough to be an insect AKHR, as opposed to the other GnRH-related receptors of insects, such as ACP and Crz receptors. The phylogenetic analyses support the data obtained from other studies involving receptors for AKH, Crz and ACP peptides. The receptor assay results with AKH analogs corroborated most of the results obtained previously using