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Proc Natl Acad Sci U S A. 2016 Jul 12;113(28):7786-91. doi: 10.1073/pnas.1608061113. Epub 2016 Jun 28.

Gpr161 anchoring of PKA consolidates GPCR and cAMP signaling.

Author information

1
Institute of Biochemistry and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria;
2
Department of Chemistry and Biochemistry, University of California, San Diego, CA 92093;
3
Institute of Molecular Biology, University of Innsbruck, 6020 Innsbruck, Austria;
4
Institute for Research in Immunology and Cancer, Université de Montréal, Montreal, QC, Canada H3C 3J7; Département de Biochimie, Université de Montréal, Montreal, QC, Canada H3C 3J7;
5
Otto-Warburg Laboratory, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany;
6
Department of Chemistry and Biochemistry, University of California, San Diego, CA 92093; Department of Pharmacology, University of California, San Diego, CA 92093;
7
Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow, G12 8QQ, United Kingdom;
8
Institute for Research in Immunology and Cancer, Université de Montréal, Montreal, QC, Canada H3C 3J7; Department of Chemistry, Université de Montréal, Montreal, QC, Canada H3C 3J7;
9
Otto-Warburg Laboratory, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany; Institute of Pharmaceutical Sciences, Pharmaceutical Chemistry, University of Graz, 8010 Graz, Austria.
10
Department of Chemistry and Biochemistry, University of California, San Diego, CA 92093; Department of Pharmacology, University of California, San Diego, CA 92093; staylor@ucsd.edu eduard.stefan@uibk.ac.at.
11
Institute of Biochemistry and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria; staylor@ucsd.edu eduard.stefan@uibk.ac.at.

Abstract

Scaffolding proteins organize the information flow from activated G protein-coupled receptors (GPCRs) to intracellular effector cascades both spatially and temporally. By this means, signaling scaffolds, such as A-kinase anchoring proteins (AKAPs), compartmentalize kinase activity and ensure substrate selectivity. Using a phosphoproteomics approach we identified a physical and functional connection between protein kinase A (PKA) and Gpr161 (an orphan GPCR) signaling. We show that Gpr161 functions as a selective high-affinity AKAP for type I PKA regulatory subunits (RI). Using cell-based reporters to map protein-protein interactions, we discovered that RI binds directly and selectively to a hydrophobic protein-protein interaction interface in the cytoplasmic carboxyl-terminal tail of Gpr161. Furthermore, our data demonstrate that a binary complex between Gpr161 and RI promotes the compartmentalization of Gpr161 to the plasma membrane. Moreover, we show that Gpr161, functioning as an AKAP, recruits PKA RI to primary cilia in zebrafish embryos. We also show that Gpr161 is a target of PKA phosphorylation, and that mutation of the PKA phosphorylation site affects ciliary receptor localization. Thus, we propose that Gpr161 is itself an AKAP and that the cAMP-sensing Gpr161:PKA complex acts as cilium-compartmentalized signalosome, a concept that now needs to be considered in the analyzing, interpreting, and pharmaceutical targeting of PKA-associated functions.

KEYWORDS:

interaction network; molecular interactions; phosphorylation; primary cilium; scaffolding function

PMID:
27357676
PMCID:
PMC4948347
DOI:
10.1073/pnas.1608061113
[Indexed for MEDLINE]
Free PMC Article

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