title: Dual-Color Luciferase Complementation for Chemokine Receptor Signaling.
abstract: Chemokine receptors may share common ligands, setting up potential competition for ligand binding, and association of activated receptors with downstream signaling molecules such as β-arrestin. Determining the "winner" of competition for shared effector molecules is essential for understanding integrated functions of chemokine receptor signaling in normal physiology, disease, and response to therapy. We describe a dual-color click beetle luciferase complementation assay for cell-based analysis of interactions of two different chemokine receptors, CXCR4 and ACKR3, with the intracellular scaffolding protein β-arrestin 2. This assay provides real-time quantification of receptor activation and signaling in response to chemokine CXCL12. More broadly, this general imaging strategy can be applied to quantify interactions of any set of two proteins that interact with a common binding partner.
authors.affiliation: Department of Radiology, Center for Molecular Imaging, University of Michigan, Ann Arbor, Michigan, USA. Electronic address: kluker@umich.edu.
authors.forename: Kathryn E
authors.initials: KE
authors.lastname: Luker
authors.affiliation: Department of Radiology, Center for Molecular Imaging, University of Michigan, Ann Arbor, Michigan, USA; Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan, USA; Department of Microbiology and Immunology, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Gary D
authors.initials: GD
authors.lastname: Luker
mesh_terms: D054377:Chemokine CXCL12 / Q000235:genetics / Q000378:metabolism; D005816:Genetic Complementation Test; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D049409:Luciferases, Firefly / Q000235:genetics / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods; D008967:Molecular Biology / Q000379:methods*; D054387:Receptors, CXCR / Q000235:genetics / Q000378:metabolism*; D019718:Receptors, CXCR4 / Q000235:genetics / Q000378:metabolism*; D019707:Receptors, Chemokine / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D000071560:beta-Arrestin 2 / Q000378:metabolism
chemical_list: C501896:ACKR3 protein, human; C527276:CXCR4 protein, human; D054377:Chemokine CXCL12; D054387:Receptors, CXCR; D019718:Receptors, CXCR4; D019707:Receptors, Chemokine; D011993:Recombinant Fusion Proteins; D000071560:beta-Arrestin 2; D008156:Luciferases; D049409:Luciferases, Firefly
keywords: Bioluminescence; Molecular imaging; Optical imaging; Protein fragment complementation; Split luciferase