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Chemogenetics defines receptor-mediated functions of short chain free fatty acids

Nature Chemical Biology volume 15, pages 489–498 (2019)Cite this article

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Abstract

Differentiating actions of short chain fatty acids (SCFAs) at free fatty acid receptor 2 (FFA2) from other free fatty acid-responsive receptors and from non-receptor-mediated effects has been challenging. Using a novel chemogenetic and knock-in strategy, whereby an engineered variant of FFA2 (FFA2-DREADD) that is unresponsive to natural SCFAs but is instead activated by sorbic acid replaced the wild-type receptor, we determined that activation of FFA2 in differentiated adipocytes and colonic crypt enteroendocrine cells of mouse accounts fully for SCFA-regulated lipolysis and release of the incretin glucagon-like peptide-1 (GLP-1), respectively. In vivo studies confirmed the specific role of FFA2 in GLP-1 release and also demonstrated a direct role for FFA2 in accelerating gut transit. Thereby, we establish the general principle that such a chemogenetic knock-in strategy can successfully define novel G-protein-coupled receptor (GPCR) biology and provide both target validation and establish therapeutic potential of a ‘hard to target’ GPCR.

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Fig. 1: G protein interaction patterns and signal regulation by specific agonists of hFFA2 and hFFA2-DREADD are indistinguishable.
Fig. 2: Initial characterization of hFFA2-DREADD-HA expressing mice.
Fig. 3: Immunocytochemical detection of expression of hFFA2-DREADD-HA.
Fig. 4: Activation of hFFA2-DREADD-HA promotes release of GLP-1.
Fig. 5: Activation of hFFA2-DREADD-HA is anti-lipolytic.
Fig. 6: In vivo activation of hFFA2-DREADD-HA reveals roles in GLP-1 release and gut transit.

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Data availability

All data is available from the corresponding authors or is available through the University of Glasgow online data repository.

Code availability

Concentration-response curves analyzed using GraphPad Prism v.5.02.

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Acknowledgements

These studies were funded by Biotechnology and Biosciences Research Council grant nos. BB/L027887/1 (Milligan) and BB/L02781X/1 (Tobin) and a CIHR Foundation grant no. FDN-148431 (Bouvier). S.J.B. is funded through a University of Glasgow Lord Kelvin Adam Smith Fellowship and an MRC project grant (no. MR/P019366/1). We are also grateful to M. Hogue and V. Lukasheva for the design, construction and characterization of the GRK2-based biosensor. We acknowledge the BSU facilities at the Cancer Research UK Beatson Institute (C596/A17196) and the Biological Services at the University of Glasgow.

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Authors and Affiliations

  1. Centre for Translational Pharmacology, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, UK

    Daniele Bolognini, Natasja Barki, Brian D. Hudson, Eugenia Sergeev, Colin Molloy, Catherine E. Moss, Sophie J. Bradley, Andrew B. Tobin & Graeme Milligan

  2. UK Dementia Research Institute at the University of Cambridge, Cambridge, UK

    Adrian J. Butcher

  3. Department of Biochemistry and Molecular Medicine, Institute for Research in Immunology and Cancer, Université de Montreal, Montreal, Quebec, Canada

    Christian Le Gouill & Michel Bouvier

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Contributions

G.M. and A.B.T. devised the program of work. G.M., A.B.T. and D.B. wrote the paper with assistance from all other authors. G.M., A.B.T., D.B. and N.B. designed the experiments. D.B. (genetic characterization, lipolysis, GLP-1 release, immunostaining, pharmacological characterization), A.J.B. (biosensor studies), E.S. (ligand binding studies), C.L.G. (biosensor studies), C.E.M. (GLP-1 release) and N.B. (GLP-1 release and gut transit) performed experiments. B.D.H. developed the hFFA2-DREADD receptor and performed initial characterization. M.B. directed the biosensor studies. S.J.B. and C.M. oversaw the animal-based studies.

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Correspondence to Andrew B. Tobin or Graeme Milligan.

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Bolognini, D., Barki, N., Butcher, A.J. et al. Chemogenetics defines receptor-mediated functions of short chain free fatty acids. Nat Chem Biol 15, 489–498 (2019). https://doi.org/10.1038/s41589-019-0270-1

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