title: Ryanodine receptor regulation by intramolecular interaction between cytoplasmic and transmembrane domains.
abstract: Ryanodine receptors (RyR) function as Ca(2+) channels that regulate Ca(2+) release from intracellular stores to control a diverse array of cellular processes. The massive cytoplasmic domain of RyR is believed to be responsible for regulating channel function. We investigated interaction between the transmembrane Ca(2+)-releasing pore and a panel of cytoplasmic domains of the human cardiac RyR in living cells. Expression of eGFP-tagged RyR constructs encoding distinct transmembrane topological models profoundly altered intracellular Ca(2+) handling and was refractory to modulation by ryanodine, FKBP12.6 and caffeine. The impact of coexpressing dsRed-tagged cytoplasmic domains of RyR2 on intracellular Ca(2+) phenotype was assessed using confocal microscopy coupled with parallel determination of in situ protein: protein interaction using fluorescence resonance energy transfer (FRET). Dynamic interactions between RyR cytoplasmic and transmembrane domains were mediated by amino acids 3722-4610 (Interacting or "I"-domain) which critically modulated intracellular Ca(2+) handling and restored RyR sensitivity to caffeine activation. These results provide compelling evidence that specific interaction between cytoplasmic and transmembrane domains is an important mechanism in the intrinsic modulation of RyR Ca(2+) release channels.
authors.affiliation: Wales Heart Research Institute, Department of Cardiology, University of Wales College of Medicine, Cardiff, United Kingdom CF14 4XN. georgech@cf.ac.uk
authors.forename: Christopher H
authors.initials: CH
authors.lastname: George
authors.forename: Hala
authors.initials: H
authors.lastname: Jundi
authors.forename: N Lowri
authors.initials: NL
authors.lastname: Thomas
authors.forename: Mark
authors.initials: M
authors.lastname: Scoote
authors.forename: Nicola
authors.initials: N
authors.lastname: Walters
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Williams
authors.forename: F Anthony
authors.initials: FA
authors.lastname: Lai
mesh_terms: D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018613:Microscopy, Confocal; D008954:Models, Biological; D017434:Protein Structure, Tertiary; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium