A Role of Corazonin Receptor in Larval-Pupal Transition and Pupariation in the Oriental Fruit Fly Bactrocera dorsalis (Hendel) (Diptera: Tephritidae)

Abstract

Corazonin (Crz) is a neuropeptide hormone, but also a neuropeptide modulator that is internally released within the CNS, and it has a widespread distribution in insects with diverse physiological functions. Here, we identified and cloned the cDNAs of <i>Bactrocera dorsalis</i> that encode Crz and its receptor CrzR. Mature <i>BdCrz</i> has 11 residues with a unique Ser¹¹ substitution (instead of the typical Asn) and a His in the evolutionary variable position 7. The <i>BdCrzR</i> cDNA encodes a putative protein of 608 amino acids with 7 putative transmembrane domains, typical for the structure of G-protein-coupled receptors. When expressed in Chinese hamster ovary (CHO) cells, the <i>BdCrzR</i> exhibited a high sensitivity and selectivity for Crz (EC₅₀ ≈ 52.5 nM). With qPCR, the developmental stage and tissue-specific expression profiles in <i>B. dorsalis</i> demonstrated that both <i>BdCrz</i> and <i>BdCrzR</i> were highly expressed in the larval stage, and <i>BdCrzR</i> peaked in 2-day-old 3rd-instar larvae, suggesting that the <i>BdCrzR</i> may play an important role in the larval-pupal transition behavior. Immunochemical localization confirmed the production of Crz in the central nervous system (CNS), specifically by a group of three neurons in the dorso-lateral protocerebrum and eight pairs of lateral neurons in the ventral nerve cord. qPCR analysis located the <i>BdCrzR</i> in both the CNS and epitracheal gland, containing the Inka cells. Importantly, dsRNA-<i>BdCrzR</i>-mediated gene-silencing caused a delay in larval-pupal transition and pupariation, and this phenomenon agreed with a delayed expression of tyrosine hydroxylase and dopa-decarboxylase genes. We speculate that CrzR-silencing blocked dopamine synthesis, resulting in the inhibition of pupariation and cuticular melanization. Finally, injection of Crz in head-ligated larvae could rescue the effects. These findings provide a new insight into the roles of Crz signaling pathway components in <i>B. dorsalis</i> and support an important role of CrzR in larval-pupal transition and pupariation behavior.