title: Redox sensitivity of the ryanodine receptor interaction with FK506-binding protein.
abstract: The ryanodine receptor (RyR) calcium release channel functions as a redox sensor that is sensitive to channel modulators. The FK506-binding protein (FKBP) is an important regulator of channel activity, and disruption of the RyR2-FKBP12.6 association has been implicated in cardiac disease. In the present study, we investigated whether the RyR-FKBP association is redox-regulated. Using co-immunoprecipitation assays of solubilized native RyR2 from cardiac muscle sarcoplasmic reticulum (SR) with recombinant [(35)S]FKBP12.6, we found that the sulfydryl-oxidizing agents, H(2)O(2) and diamide, result in diminished RyR2-FKBP12.6 binding. Co-sedimentation experiments of cardiac SR vesicles with [(35)S]FKBP12.6 also demonstrated that oxidizing reagents decreased FKBP binding. Matching results were obtained with skeletal muscle SR. Notably, H(2)O(2) and diamide differentially affected the RyR2-FKBP12.6 interaction, decreasing binding to approximately 75 and approximately 50% of control, respectively. In addition, the effect of H(2)O(2) was negligible when the channel was in its closed state or when applied after FKBP binding had occurred, whereas diamide was always effective. A cysteine-null mutant FKBP12.6 retained redox-sensitive interaction with RyR2, suggesting that the effect of the redox reagents is exclusively via sites on the ryanodine receptor. K201 (or JTV519), a drug that has been proposed to prevent FKBP12.6 dissociation from the RyR2 channel complex, did not restore normal FKBP binding under oxidizing conditions. Our results indicate that the redox state of the RyR is intimately connected with FKBP binding affinity.
authors.affiliation: Wales Heart Research Institute, Department of Cardiology, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom. zissimopouloss@cardiff.ac.uk
authors.forename: Spyros
authors.initials: S
authors.lastname: Zissimopoulos
authors.forename: Naadiya
authors.initials: N
authors.lastname: Docrat
authors.forename: F Anthony
authors.initials: FA
authors.lastname: Lai
mesh_terms: D002118:Calcium / Q000378:metabolism; D003958:Diamide / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D010084:Oxidation-Reduction; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology*; D013481:Superoxides / Q000378:metabolism; D022021:Tacrolimus Binding Proteins / Q000502:physiology*
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D003958:Diamide; D013481:Superoxides; D006861:Hydrogen Peroxide; D022021:Tacrolimus Binding Proteins; C403484:tacrolimus binding protein 1B; D005978:Glutathione; D002118:Calcium; D004229:Dithiothreitol