title: Caveolae-specific activation loop between CaMKII and L-type Ca2+channel aggravates cardiac hypertrophy in α1-adrenergic stimulation.
abstract: Activation of CaMKII induces a myriad of biological processes and plays dominant roles in cardiac hypertrophy. Caveolar microdomain contains many calcium/calmodulin-dependent kinase II (CaMKII) targets, including L-type Ca2+channel (LTCC) complex, and serves as a signaling platform. The location of CaMKII activation is thought to be critical; however, the roles of CaMKII in caveolae are still elusive due to lack of methodology for the assessment of caveolae-specific activation. Our aim was to develop a novel tool for the specific analysis of CaMKII activation in caveolae and to determine the functional role of caveolar CaMKII in cardiac hypertrophy. To assess the caveolae-specific activation of CaMKII, we generated a fusion protein composed of phospholamban and caveolin-3 (cPLN-Cav3) and GFP fusion protein with caveolin-binding domain fused to CaMKII inhibitory peptide (CBD-GFP-AIP), which inhibits CaMKII activation specifically in caveolae. Caveolae-specific activation of CaMKII was detected using phosphospecific antibody for PLN (Thr17). Furthermore, adenoviral overexpression of LTCC β2a-subunit (β2a) in NRCMs showed its constitutive phosphorylation by CaMKII, which induces hypertrophy, and that both phosphorylation and hypertrophy are abolished by CBD-GFP-AIP expression, indicating that β2aphosphorylation occurs specifically in caveolae. Finally, β2aphosphorylation was observed after phenylephrine stimulation in β2a-overexpressing mice, and attenuation of cardiac hypertrophy after chronic phenylephrine stimulation was observed in nonphosphorylated mutant of β2a-overexpressing mice. We developed novel tools for the evaluation and inhibition of caveolae-specific activation of CaMKII. We demonstrated that phosphorylated β2adominantly localizes to caveolae and induces cardiac hypertrophy after α1-adrenergic stimulation in mice.NEW & NOTEWORTHYWhile signaling in caveolae is thought to be important in cardiac hypertrophy, direct evidence is missing due to lack of tools to assess caveolae-specific signaling. This is the first study to demonstrate caveolae-specific activation of CaMKII signaling in cardiac hypertrophy induced by α1-adrenergic stimulation using an originally developed tool.
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Kota
authors.initials: K
authors.lastname: Tonegawa
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Otsuka
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Shohei
authors.initials: S
authors.lastname: Kumagai
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Sachi
authors.initials: S
authors.lastname: Matsunami
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Nao
authors.initials: N
authors.lastname: Hayamizu
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Shota
authors.initials: S
authors.lastname: Tanaka
authors.affiliation: Faculty of Medicine, Department of Pharmacology, Osaka Medical College, Takatsuki, Osaka, Japan; and.
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Moriwaki
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Masanori
authors.initials: M
authors.lastname: Obana
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Makiko
authors.initials: M
authors.lastname: Maeda
authors.affiliation: Faculty of Medicine, Department of Pharmacology, Osaka Medical College, Takatsuki, Osaka, Japan; and.
authors.forename: Michio
authors.initials: M
authors.lastname: Asahi
authors.affiliation: Animal Resource Development Unit and Genetic Engineering Team, RIKEN Center for Life Science Technologies, Kobe, Hyogo, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kiyonari
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Fujio
authors.affiliation: Laboratory of Clinical Science and Biomedicine, Graduate School of Pharmacological Sciences, Osaka University, Suita, Osaka, Japan; nakayama@phs.osaka-u.ac.jp.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nakayama
mesh_terms: D058646:Adrenergic alpha-1 Receptor Agonists; D000818:Animals; D000831:Animals, Newborn; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000235:genetics / Q000378:metabolism*; D006332:Cardiomegaly / Q000139:chemically induced / Q000201:enzymology / Q000378:metabolism*; D021941:Caveolae / Q000201:enzymology / Q000378:metabolism*; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D005355:Fibrosis; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D051381:Rats; D014162:Transfection
chemical_list: D058646:Adrenergic alpha-1 Receptor Agonists; D020746:Calcium Channels, L-Type; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2
keywords: L-type calcium channel; calcium/calmodulin dependent kinase II; caveolae microdomain