title: New insights into the model of dopamine D1 receptor and G-proteins interactions.
abstract: The details of the interaction between G-proteins and the GPCRs have been subjected to extensive investigation with structural and functional assays, but still many fundamental questions regarding this macromolecular assembly and its mechanism remain unanswered. In the context of current structural data we investigated interactions of dopamine D1 receptor with cognate G-proteins (Gαs) in living cells, emphasizing the prevalence of preassembled D1-G-protein complexes. We also tested the effect of D1 receptor presence on the dynamics of Gαs and Gαi3 in the cellular plasma membrane. Using fluorescence resonance energy transfer (FRET) detected by fluorescence lifetime imaging microscopy (FLIM) or fluorescence recovery after photobleaching (FRAP) microscopy, we did not detect constitutive preassociated complex between D1 receptor and G-protein in the absence of receptor activation. Our work suggests that D1 receptor alters the distribution of Gαs and Gαi3 subunits inside the membrane. We also find that non-activated D1 receptor and Gαs or Gαi3 are present in the cell membrane within the same membrane microdomains in the proximity of about 9-10 nm.
authors.affiliation: Department of Physical Biochemistry, Faculty of Biochemistry Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland.
authors.forename: Paweł
authors.initials: P
authors.lastname: Mystek
authors.affiliation: Department of Physical Biochemistry, Faculty of Biochemistry Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Tworzydło
authors.affiliation: Department of Physical Biochemistry, Faculty of Biochemistry Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland.
authors.forename: Marta
authors.initials: M
authors.lastname: Dziedzicka-Wasylewska
authors.affiliation: Department of Physical Biochemistry, Faculty of Biochemistry Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland. Electronic address: a.polit@uj.edu.pl.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Polit
mesh_terms: D001552:Benzazepines / Q000494:pharmacology; D002839:Chromans / Q000494:pharmacology; D031541:Fluorescence Resonance Energy Transfer; D019204:GTP-Binding Proteins / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D018613:Microscopy, Confocal; D008954:Models, Biological; D011485:Protein Binding / Q000187:drug effects; D017447:Receptors, Dopamine D1 / Q000819:agonists / Q000378:metabolism*; D014018:Tissue Distribution
chemical_list: D001552:Benzazepines; D002839:Chromans; D017447:Receptors, Dopamine D1; C067113:SK&F 81297; C065380:A 68930; D019204:GTP-Binding Proteins
keywords: Dopamine receptor; FLIM–FRET; FRAP; G-protein; G-protein coupling; GPCR