Molecular Characterization and Functional Analysis of a Putative Octopamine/Tyramine Receptor during the Developmental Stages of the Pacific Oyster, Crassostrea gigas

PLoS One. 2016 Dec 16;11(12):e0168574. doi: 10.1371/journal.pone.0168574. eCollection 2016.

Abstract

Octopamine (OA) and its precursor, tyramine (TA), participate in invertebrate development such as growth, maturation, and reproduction by activating their corresponding G protein-coupled receptors (GPCRs). Although OA was first discovered in mollusks (octopus), subsequent studies on OA, TA and related receptors have primarily been conducted in Ecdysozoa, especially in insects. Accordingly, only limited reports on OA/TA receptors in mollusks are available and their physiological roles remain unclear. Here, a full-length cDNA encoding a putative 524 amino acid OA/TA receptor (CgGPR1) was isolated from the Pacific oyster Crassostrea gigas. CgGPR1 was most closely related to the Lymnaea stagnalis OA receptor OAR2 in sequence. Phylogenetic analysis showed that CgGPR1 belongs to a poorly studied subfamily of invertebrate OA/TA receptors. The spatio-temporal expression of CgGPR1 in C. gigas larvae was examined by quantitative real-time PCR and Western blot analysis. CgGPR1 was expressed during all developmental stages of C. gigas with higher levels at mid-developmental stages, indicating its potential role in embryogenesis and tissue differentiation. Immunoreactive fluorescence of CgGPR1 was mainly observed in the velum, foot, gill and mantle of C. gigas larvae. CgGPR1 transcripts were detected in all the tested organs of adult C. gigas, with highest level in the mantle. Pharmacological analysis showed that cAMP and Ca2+ concentrations remained unchanged in HEK293 cells expressing CgGPR1 upon addition of OA, TA or related amines, suggesting that CgGPR1 modulates other unknown molecules rather than cAMP and Ca2+. Our study sheds light on CgGPR1 function in oysters.

MeSH terms

  • Animals
  • Calcium / metabolism
  • Cloning, Molecular / methods*
  • Cyclic AMP / metabolism
  • Gene Expression Regulation, Developmental
  • HEK293 Cells
  • Humans
  • Ostreidae / growth & development*
  • Ostreidae / metabolism
  • Phylogeny
  • Receptors, Biogenic Amine / genetics
  • Receptors, Biogenic Amine / metabolism
  • Receptors, G-Protein-Coupled / genetics*
  • Receptors, G-Protein-Coupled / metabolism*

Substances

  • Receptors, Biogenic Amine
  • Receptors, G-Protein-Coupled
  • norsynephrine receptor
  • tyramine receptor
  • Cyclic AMP
  • Calcium

Grants and funding

This research was supported by the National Natural Science Foundation of China (31402285, 31530079), the National Basic Research Program of China (973 Program, 2010CB126401), the Earmarked Fund for Modern Agro-industry Technology Research System (CARS-48), the Scientific and Technological Innovation Project Financially Supported by Qingdao National Laboratory for Marine Science and Technology (2015ASKJ02-03), and Taishan Scholars Climbing Program of Shandong. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.