title: Suppression of Rad leads to arrhythmogenesis via PKA-mediated phosphorylation of ryanodine receptor activity in the heart.
abstract: Ras-related small G-protein Rad plays a critical role in generating arrhythmias via regulation of the L-type Ca(2+) channel (LTCC). The aim was to demonstrate the role of Rad in intracellular calcium homeostasis by cardiac-Specific dominant-negative suppression of Rad. Transgenic (TG) mice overexpressing dominant-negative mutant Rad (S105N Rad TG) were generated. To measure intracellular Ca(2+) concentration ([Ca(2+)]i), we recorded [Ca(2+)]i transients and Ca(2+) sparks from isolated cardiomyocytes using confocal microscopy. The mean [Ca(2+)]i transient amplitude was significantly increased in S105N Rad TG cardiomyocytes, compared with control littermate mouse cells. The frequency of Ca(2+) sparks was also significantly higher in TG cells than in control cells, although there were no significant differences in amplitude. The sarcoplasmic reticulum Ca(2+) content was not altered in the S105N Rad TG cells, as assessed by measuring caffeine-induced [Ca(2+)]i transient. In contrast, phosphorylation of Ser(2809) on the cardiac ryanodine receptor (RyR2) was significantly enhanced in TG mouse hearts compared with controls. Additionally, the Rad-mediated RyR2 phosphorylation was regulated via a direct interaction of Rad with protein kinase A (PKA).
authors.affiliation: Department of Cardiology, School of Medicine, Keio University, Tokyo, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamakawa
authors.affiliation: Department of Laboratory Medicine, School of Medicine, Keio University, Tokyo, Japan. Electronic address: muratam@a7.keio.jp.
authors.forename: Mitsushige
authors.initials: M
authors.lastname: Murata
authors.affiliation: Department of Cardiovascular Research, Research Institute of Environmental Medicine, Nagoya University, Nagoya, Japan.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Suzuki
authors.affiliation: Department of Cardiology, First Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Hirotaka
authors.initials: H
authors.lastname: Yada
authors.affiliation: Department of Physiology, Tokai University School of Medicine, Kanagawa, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Ishida
authors.affiliation: Department of Cardiology, School of Medicine, Keio University, Tokyo, Japan.
authors.forename: Yoshiyasu
authors.initials: Y
authors.lastname: Aizawa
authors.affiliation: Department of Cardiology, First Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Adachi
authors.affiliation: Department of Cardiovascular Research, Research Institute of Environmental Medicine, Nagoya University, Nagoya, Japan.
authors.forename: Kaichiro
authors.initials: K
authors.lastname: Kamiya
authors.affiliation: Department of Cardiology, School of Medicine, Keio University, Tokyo, Japan.
authors.forename: Keiichi
authors.initials: K
authors.lastname: Fukuda
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001145:Arrhythmias, Cardiac / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D009206:Myocardium / Q000378:metabolism / Q000473:pathology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation; D061251:Primary Cell Culture; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D018631:ras Proteins / Q000172:deficiency / Q000235:genetics*
chemical_list: C490030:Rrad protein, mouse; D019837:Ryanodine Receptor Calcium Release Channel; C000592748:ryanodine receptor 2. mouse; D002110:Caffeine; D017868:Cyclic AMP-Dependent Protein Kinases; D018631:ras Proteins; D002118:Calcium
keywords: Ca(2+) imaging; Excitation–contraction (EC) coupling (ECC); PKA signaling; Rad (Ras associated with diabetes); Ryanodine receptor