Prokineticin receptors interact unselectively with several G protein subtypes but bind selectively to β-arrestin 2 |
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| Institution: | 1. IRCCS Fondazione Santa Lucia, Via del Fosso di Fiorano 64, 00143 Rome, Italy;2. Institute of Cell Biology and Neurobiology, CNR, Via del Fosso di Fiorano, 64, 00143 Rome, Italy;3. European Brain Research Institute, Via del Fosso di Fiorano, 64, 00143 Rome, Italy;4. Department of Life and Environmental Sciences, University of Cagliari, Via Ospedale, 72, 09124 Cagliari, Italy;5. Department of Experimental Medicine and Surgery, University of Rome “Tor Vergata”, Via Montpellier, 1, 00144 Rome, Italy;6. Department of Human Physiology and Pharmacology “Vittorio Erspamer”, University of Roma “La Sapienza”, P.za A. Moro 5, 00185 Rome, Italy;7. Department of Biology, University of Rome “Tor Vergata”, Via della Ricerca Scientifica, 1, 00133 Rome, Italy;8. Department of Systems Medicine, University of Rome “Tor Vergata”, Via Montpellier, 1, 00133 Rome, Italy;1. Institut national de la santé et de la recherche médicale, unité 1036, University Grenoble-Alpes, 38041 Grenoble, France;2. Commissariat à l’énergie atomique (CEA), BIG-Biology of Cancer and Infection, 38054 Grenoble, France;3. State Key Laboratory of Medical Genetics, Central South University, Changsha, Hunan 410078, China,;4. Department of Basic Medical Sciences, Changsha Medical University, Changsha, Hunan 410219, China,;6. Department of Pharmacology, Hubei University of Science and Technology, 88 Xianning Road, Xianning, Hubei 437100, China, and;5. Department of Pharmacology, University of California, Irvine, California 92697 |
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| Abstract: | Prokineticin 1 (pk1) and prokineticin 2 (pk2) interact with two structurally related G-protein coupled receptors, prokineticin receptor 1 (PKR1) and prokineticin receptor 2 (PKR2). Cellular signalling studies show that the activated receptors can evoke Ca2+-mobilization, pertussis toxin-sensitive ERK phosphorylation, and intracellular cAMP accumulation, which suggests the partecipation of several G protein subtypes, such as Gq/11, Gi/o and Gs. However, direct interactions with these transduction proteins have not been studied yet. Here we measured by bioluminescence resonance energy transfer (BRET) the association of PKR1 and PKR2 with different heterotrimeric Gα proteins in response to pk1 and pk2 activation. Using host-cell lines carrying gene deletions of Gαq/11 or Gαs, and pertussis toxin treatment to abolish the receptor interactions with Gαi/o, we determined that both receptors could couple with comparable efficiency to Gq/11 and Gi/o, but far less efficiently to Gs or other pertussis toxin-insensitive G proteins. We also used BRET methodology to assess the association of prokineticin receptors with β-arrestin isoforms. Fluorescent versions of the isoforms were transfected both in HEK293 cells and in double KO β-arrestin 1/2 mouse fibroblasts, to study receptor interaction with the reconstituted individual β-arrestins without background expression of the endogenous genes. Both receptors formed stable BRET-emitting complexes with β-arrestin 2 but not with β-arrestin 1, indicating strong selectivity for the former. In all the studied transducer interactions and in both receptors, pk2 was more potent than pk1 in promoting receptor binding to transduction proteins. |
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