1. Academic Validation
  2. Rho kinase-dependent desensitization of GPR39; a unique mechanism of GPCR downregulation

Rho kinase-dependent desensitization of GPR39; a unique mechanism of GPCR downregulation

  • Biochem Pharmacol. 2017 Sep 15;140:105-114. doi: 10.1016/j.bcp.2017.06.115.
Yuji Shimizu 1 Ryokichi Koyama 2 Tomohiro Kawamoto 2
Affiliations

Affiliations

  • 1 Biomolecular Research Laboratories, Pharmaceutical Research Division, Takeda Pharmaceutical Company Limited, 26-1, Muraoka-higashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan. Electronic address: yuuji.shimizu@takeda.com.
  • 2 Biomolecular Research Laboratories, Pharmaceutical Research Division, Takeda Pharmaceutical Company Limited, 26-1, Muraoka-higashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan.
Abstract

GPR39, a G-protein-coupled receptor activated by zinc, reportedly activates multiple intracellular signaling pathways via Gs, Gq, G12/13, and β-arrestin, but little is known about downregulation of the receptor upon its activation. To our knowledge, this is the first report on the mechanism of feedback regulation of GPR39 function determined in GPR39-expressing HEK293 cells (HEK293-GPR39) as a model cell system. In HEK293-GPR39 cells, GPR39-C3, which is a positive allosteric modulator, activated cAMP production (downstream of Gs), IP1 accumulation (downstream of Gq), SRF-RE-dependent transcription (downstream of G12/13), and β-arrestin recruitment. GPR39-C3 induced dose- and time-dependent loss of response in cAMP production by second challenge of the compound. This functional desensitization was blocked by the Rho kinase (ROCK) inhibitor, Y-27632, but not by Gq or Gs-pathway inhibitors or inhibition of β-arrestin recruitment. In the receptor localization assay, GPR39-C3 induced internalization of GFP-tagged GPR39. This internalization was also inhibited by Y-27632, which suggested that ROCK activation is critical for internalization and desensitization of GPR39. A novel biased GPR39 positive allosteric modulator, 5-[2-[(2,4-dichlorophenyl)methoxy]phenyl]-2,2-dimethyl-1,3,5,6-tetrahydrobenzo[a]phenanthridin-4-one (GSB-118), which activated cAMP responses and β-arrestin recruitment but showed no effect on SRF-RE-dependent transcription, did not induce desensitization. These results revealed a unique mechanism of desensitization of GPR39.

Keywords

Biased ligand; Desensitization; G-protein-coupled receptor 39; GPR39-C3 (PubChem CID: 091826086); GSB-118 (PubChem CID: 003112012); Positive allosteric modulator; Rho kinase.

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