Development and Reproduction (한국발생생물학회지:발생과생식)

The Korean Society of Developmental Biology (한국발생생물학회)
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The C-terminal Phosphorylation Sites of eel Follicle-Stimulating Hormone Receptor are Important Role in the Signal Transduction

  • Kim, Jeong-Min (Animal Biotechnology, Graduate School of Future Convergence Technology, Dept. of Animal Life Science, Institute of Genetic Engineering, Hankyong National University) ;
  • Byambaragchaa, Munkhzaya (Animal Biotechnology, Graduate School of Future Convergence Technology, Dept. of Animal Life Science, Institute of Genetic Engineering, Hankyong National University) ;
  • Kang, Myung-Hwa (Dept. of Food Science and Nutrition, Hoseo University) ;
  • Min, Kwan-Sik (Animal Biotechnology, Graduate School of Future Convergence Technology, Dept. of Animal Life Science, Institute of Genetic Engineering, Hankyong National University)
  • Received : 2018.06.09
  • Accepted : 2018.06.22
  • Published : 2018.06.30
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Abstract

The large extracellular domain of glycoprotein hormone receptors is a unique feature within the G protein-coupled receptors (GPCRs) family. After interaction with the hormone, the receptor becomes coupled to Gs, which, in turn stimulates adenylyl cyclase and the production of cAMP. Potential phosphorylation sites exist in the C-terminal region of GPCRs. The experiments described herein represent attempts to determine the functions of the eel follicle-stimulating hormone receptor (eelFSHR). We constructed a mutant of eelFSHR, in which the C-terminal cytoplasmic tail was truncated at residue 614 (eelFSHR-t614). The eelFSHR-t614 lacked all potential phosphorylation sites present in the C-terminal region of eelFSHR. In order to obtain the eelFSHR ligand, we produced recombinant follicle-stimulating hormone ($rec-eelFSH{\beta}/{\alpha}$) in the CHO-suspension cells. The expression level was 2-3 times higher than that of the transient expression of eelFSH in attached CHO-K1 cells. The molecular weight of the $rec-eelFSH{\beta}/{\alpha}$ protein was identified to be approximately 34 kDa. The cells expressing eelFSHR-t614 showed an increase in agonist-induced cAMP responsiveness. The maximal cAMP responses of cells expressing eelFSHR-t614 were lower than those of cells expressing eelFSHR-wild type (eelFSHR-WT). The $EC_{50}$ following C-terminal deletion in CHO-K1 cells was approximately 60.4% of that of eelFSHR-WT. The maximal response in eelFSHR-t614 cells was also drastically lower than that of eelFSHR-WT. We also found similar results in PathHunter Parental cells expressing ${\beta}$-arrestin. Thus, these data provide evidence that the truncation of the C-terminal cytoplasmic tail phosphorylation sites in the eelFSHR greatly decreased cAMP responsiveness and maximal response in both CHO-K1 cells and Path-Hunter Parental cells expressing ${\beta}$-arrestin.

Keywords

References

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