Specific Activation of the G Protein-coupled Receptor BNGR-A21 by the Neuropeptide Corazonin from the Silkworm, Bombyx mori, Dually Couples to the Gq and Gs Signaling Cascades

Jingwen Yang,Haishan Huang,Huipeng Yang,Xiaobai He,Xue Jiang,Ying Shi,Damirin Alatangaole,Liangen Shi,Naiming Zhou

doi:10.1074/jbc.m112.441675

Jingwen Yang, Haishan Huang + Show 7 more

Open Access

https://doi.org/10.1074/jbc.m112.441675

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Abstract

Corazonin, an undecapeptide neurohormone sharing a highly conserved amino acid sequence across Insecta, plays different physiological roles in the regulation of heart contraction rates, silk spinning rates, the induction of dark color and morphometric phase changes, and ecdysis. Corazonin receptors have been identified in Drosophila melanogaster, Manduca sexta, and Musca domestica. However, detailed information on the signaling and major physiological functions of corazonin and its receptor is largely unknown. In the current study, using both the mammalian cell line HEK293 and insect cell lines BmN and Sf21, we paired the Bombyx corazonin neuropeptide as a specific endogenous ligand for the Bombyx neuropeptide G protein-coupled receptor A21 (BNGR-A21), and we therefore designated this receptor as BmCrzR. Further characterization indicated that synthetic BmCrz demonstrated a high affinity for and activated BmCrzR, resulting in intracellular cAMP accumulation, Ca(2+) mobilization, and ERK1/2 phosphorylation via the Gq- and Gs-coupled signaling pathways. The direct interaction of BmCrzR with BmCrz was confirmed by a rhodamine-labeled BmCrz peptide. Moreover, experiments with double-stranded RNA and synthetic peptide injection suggested a possible role of BmCrz/BmCrzR in the regulation of larval growth and spinning rate. Our present results provide the first in-depth information on BmCrzR-mediated signaling for further elucidation of the BmCrz/BmCrzR system in the regulation of fundamental physiological processes.

Highlights

The corazonin system is of importance for insects to control multiple physiological processes
Activation of the BNGR-A21 Receptor by the BmCrz Peptide Dually Couples to the Gq and Gs Signaling Pathways—Previous studies have shown that the corazonin receptor was only activated by its endogenous corazonin, leading to cAMP accumulation, in A. gambiae [22], the detailed signaling pathways remain to be elucidated
To examine BNGR-A21mediated G protein coupling and signaling, human embryonic kidney 293 (HEK293) and Sf21 cells were stably cotransfected with BNGR-A21 and a reporter gene system consisting of the firefly luciferase coding region under control of a minimal promoter containing cAMP-response elements

Summary

Background

The corazonin system is of importance for insects to control multiple physiological processes. Results: Bombyx BNGR-A21 was activated in signaling and internalization by direct interaction with corazonin. Significance: Our findings will lead to a better understanding of the corazonin system in regulation of fundamental physiological processes. Detailed information on the signaling and major physiological functions of corazonin and its receptor is largely unknown. In the current study, using both the mammalian cell line HEK293 and insect cell lines BmN and Sf21, we paired the Bombyx corazonin neuropeptide as a specific endogenous ligand for the Bombyx neuropeptide G proteincoupled receptor A21 (BNGR-A21), and we designated this receptor as BmCrzR. Our present results provide the first in-depth information on BmCrzR-mediated signaling for further elucidation of the BmCrz/BmCrzR system in the regulation of fundamental physiological processes.

The abbreviations used are

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION