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Front. Immunol. | doi: 10.3389/fimmu.2018.00415

The phosphorylation of CCR6 on distinct Ser/Thr residues in the carboxyl terminus differentially regulates biological function

Mei-Yi Lu1,  Syuan-Shao Lu1, Shiann-Luen Chang1 and  Fang Liao1*
  • 1Institute of Biomedical Sciences, Academia Sinica, Taiwan

CCR6 is a G protein-coupled receptor (GPCR) that recognizes a single chemokine ligand, CCL20, and is primarily expressed by leukocytes. Upon ligand binding, CCR6 activates Gαi heterotrimeric G proteins to induce various potential cellular outcomes through context-specific cell signaling. It is well known that differential phosphorylation of Ser and Thr residues in the C-terminal domains or intracellular loops of GPCRs can generate barcodes that regulate GPCR function by regulating the recruitment of β-arrestins. In this study, we demonstrate that ligand binding to CCR6 induces receptor phosphorylation at Ser/Thr residues in the C-terminal tail, rather than intracellular loops. Using mutagenesis experiments, we determined that distinct clusters of Ser/Thr residues in the C-terminal domain differentially regulate CCL20-induced signaling and cellular response. Substituting the Thr360/Ser361/Thr363 cluster or the Ser370/Ser371 cluster with Ala residues modulated cellular response upon CCL20 stimulation. Notably, receptor internalization, chemotaxis, F-actin distribution, transient ERK1/2 activation and β-arrestin 2 recruitment were oppositely affected by mutating the two clusters, suggesting that phosphorylation of CCR6 C-terminal Ser/Thr residues directs the cell signaling response upon receptor activation. Moreover, activated CCR6 weakly recruited β-arrestin 1 in comparison to β-arrestin 2, and the two arrestin proteins seemed to play overlapping but distinct roles in mediating CCL20/CCR6-induced cellular responses. Taken together, the effects of site-specific Ser/Thr phosphorylation on CCR6 demonstrate the existence of barcodes on the protein that dictate the activation of different cell signaling profiles and lead to distinct biological outcomes.

Keywords: CCR6, GPCR, Mutagenesis, receptor phosphorylation, beta-arrestins

Received: 22 Dec 2017; Accepted: 15 Feb 2018.

Edited by:

Mette M. Rosenkilde, University of Copenhagen, Denmark

Reviewed by:

Jose Miguel Rodriguez Frade, Consejo Superior de Investigaciones Científicas (CSIC), Spain
José Luis Rodríguez-Fernández, Consejo Superior de Investigaciones Científicas (CSIC), Spain  

Copyright: © 2018 Lu, Lu, Chang and Liao. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Dr. Fang Liao, Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan,