Concomitant action of structural elements and receptor phosphorylation determine arrestin-3 interaction with the free fatty acid receptor FFA4

Butcher, A. J., Hudson, B. D. , Shimpukade, B., Alvarez-Curto, E. , Prihandoko, R., Ulven, T., Milligan, G. and Tobin, A. B. (2014) Concomitant action of structural elements and receptor phosphorylation determine arrestin-3 interaction with the free fatty acid receptor FFA4. Journal of Biological Chemistry, 289, pp. 18451-18465. (doi: 10.1074/jbc.M114.568816) (PMID:24817122) (PMCID:PMC4140278)

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In addition to being nutrients, free fatty acids also act as signalling molecules by activating a family of G protein-coupled receptors. Among these is FFA4, previously called GPR120, which responds to medium and long chain fatty acids, including health promoting omega-3 fatty acids, which have been implicated in the regulation of metabolic and inflammatory responses. Here we show, using mass spectrometry, mutagenesis and phospho-specific antibodies, that agonist regulated phosphorylation of the human FFA4 receptor occurs primarily at five residues (T347 T349 S350 S357 S360) in the C terminal tail. Mutation of these residues reduces both the efficacy and potency of ligand-mediated arrestin-3 recruitment, as well as affecting recruitment kinetics. Combined mutagenesis of all five of these residues was insufficient to fully abrogate interaction with arrestin-3 but further mutagenesis of negatively charged residues revealed additional structural components for the interaction with arrestin-3 within the C terminal tail of the receptor. These elements consist of the acidic residues E341, D348 and D355, located close to the phosphorylation sites. Receptor phosphorylation thus operates in concert with structural elements within the C-terminal tail of FFA4 to allow for the recruitment of arrestin-3. Importantly, these mechanisms of arrestin-3 recruitment operate independently from Gq/11-coupling thereby offering the possibility that ligands showing stimulus bias could be developed that exploit these differential coupling mechanisms. Furthermore, this provides a strategy for the design of biased receptors to probe physiologically relevant signalling.

Item Type:Articles
Glasgow Author(s) Enlighten ID:Alvarez-Curto, Dr Elisa and Hudson, Dr Brian and Milligan, Professor Graeme and Tobin, Andrew
Authors: Butcher, A. J., Hudson, B. D., Shimpukade, B., Alvarez-Curto, E., Prihandoko, R., Ulven, T., Milligan, G., and Tobin, A. B.
College/School:College of Medical Veterinary and Life Sciences > School of Molecular Biosciences
Journal Name:Journal of Biological Chemistry
Journal Abbr.:J Biol Chem.
Publisher:American Society for Biochemistry and Molecular Biology, Inc.
ISSN (Online):1083-351X
Copyright Holders:Copyright © 2014 The Authors
First Published:First published in Journal of Biological Chemistry 289:18451-18465
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
618921GPR120: a G protein-coupled receptor with the potential to regulate insulin secretion and inflammationGraeme MilliganBiotechnology and Biological Sciences Research Council (BBSRC)BB/K019856/1RI MOLECULAR CELL & SYSTEMS BIOLOGY