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Conditional expression and signaling of a specifically designed Gi-coupled receptor in transgenic mice

Nature Biotechnology volume 17, pages 165–169 (1999)Cite this article

Abstract

To control G protein signaling in vivo, we have modified G protein–coupled receptors to respond exclusively to synthetic small molecule agonists and not to their natural agonist(s). These engineered receptors are designated RASSLs (receptor activated solely by a synthetic ligand). A prototype RASSL (Ro1) based on the Gi–coupled κ opioid receptor was expressed in transgenic mice under the control of the tetracycline transactivator (tet) system. Activation of Ro1 expressed in the heart decreased heart rate by up to 80%, an expected effect of increased Gi signaling. Maximal heart rate changes occurred in less than 1 min, demonstrating the speed of this inducible signaling system. This Ro1–mediated slowing of heart rate was also subject to desensitization, which lasted more than 24 h. Both the initial effect on heart rate and the desensitization occurred, even though Ro1 is derived from a human opioid receptor not normally involved in heart rate control. In addition, the tet system was used to induce Ro1 expression in hepatocytes and salivary gland, where Gi signaling is known to control physiologic events such as proliferation and secretion. These studies demonstrate that a RASSL can be inducibly expressed in several mouse tissues and used in vivo to activate G protein signaling in a controllable fashion.

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Figure 1: Conditional and tissue–specific expression of Ro1 in heart, liver, and salivary gland.
Figure 2: Timing of Ro1 expression in the heart.
Figure 3: Inducible β–gal activity in the atria and ventricles of αMHC–tTA/tetO–Ro1/tetO–lacZ mice.
Figure 4: Spiradoline–mediated bradycardia in mice with cardiac Ro1 expression.
Figure 5: (A) Spiradoline–mediated bradycardia occurs immediately.
Figure 6: Average decrease in heart rate for five mice (n = 4 at 4 h) treated with spiradoline at 4, 28, and 168 h after initial injection (1 × 10–5 mol/kg).

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Acknowledgements

We thank Robert W. Mahley and Murielle M. Veniant for valuable discussions and advice; Lara Jensen and Dale Newland for histochemistry assistance; and Gary Howard, Stephen Ordway, and Jane Peredo for editorial assistance. G. Fishman is an established investigator of the American Heart Association. This work was supported by the J. David Gladstone Institutes and the National Institutes of Health Grants CA71779 (C.H.R.) and HL60664–01 (B.R.C.).

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Authors and Affiliations

  1. Gladstone Institute of Cardiovascular Disease, Gladstone Institute of Neurological Disease, University of California, San Francisco, 94141, CA

    Charles H. Redfern, Peter Coward, Michael Y. Degtyarev, Elena K. Lee, Andrew T. Kwa & Bruce R. Conklin

  2. Department of Medicine, University of California, San Francisco, 94141, CA

    Charles H. Redfern & Bruce R. Conklin

  3. National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, 20982, MD

    Lothar Hennighausen

  4. Zentrum für Moleculare Biologie der Universitat Heidelberg, Heidelberg, Germany

    Hermann Bujard

  5. Mt. Sinai School of Medicine, New York, 10029, NY

    Glenn I. Fishman

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  1. Charles H. Redfern
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Correspondence to Bruce R. Conklin.

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Redfern, C., Coward, P., Degtyarev, M. et al. Conditional expression and signaling of a specifically designed Gi-coupled receptor in transgenic mice. Nat Biotechnol 17, 165–169 (1999). https://doi.org/10.1038/6165

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