RyR2 Serine-2030 PKA Site Governs Ca2+ Release Termination and Ca2+ Alternans
Por:
Wei, JH, Guo, WT, Wang, RW, Estillore, JP, Belke, D, Chen, YX, Vallmitjana, A, Benitez, R, Hove-Madsen, L, Chen, SRW
Publicada:
20 ene 2023
Ahead of Print:
30 dic 2022
Resumen:
Background:PKA (protein kinase A)-mediated phosphorylation of cardiac RyR2 (ryanodine receptor 2) has been extensively studied for decades, but the physiological significance of PKA phosphorylation of RyR2 remains poorly understood. Recent determination of high-resolution 3-dimensional structure of RyR2 in complex with CaM (calmodulin) reveals that the major PKA phosphorylation site in RyR2, serine-2030 (S2030), is located within a structural pathway of CaM-dependent inactivation of RyR2. This novel structural insight points to a possible role of PKA phosphorylation of RyR2 in CaM-dependent inactivation of RyR2, which underlies the termination of Ca2+ release and induction of cardiac Ca2+ alternans. Methods:We performed single-cell endoplasmic reticulum Ca2+ imaging to assess the impact of S2030 mutations on Ca2+ release termination in human embryonic kidney 293 cells. Here we determined the role of the PKA site RyR2-S2030 in a physiological setting, we generated a novel mouse model harboring the S2030L mutation and carried out confocal Ca2+ imaging. Results:We found that mutations, S2030D, S2030G, S2030L, S2030V, and S2030W reduced the endoplasmic reticulum luminal Ca2+ level at which Ca2+ release terminates (the termination threshold), whereas S2030P and S2030R increased the termination threshold. S2030A and S2030T had no significant impact on release termination. Furthermore, CaM-wild-type increased, whereas Ca2+ binding deficient CaM mutant (CaM-M [a loss-of-function CaM mutation with all 4 EF-hand motifs mutated]), PKA, and Ca2+/CaMKII (CaM-dependent protein kinase II) reduced the termination threshold. The S2030L mutation abolished the actions of CaM-wild-type, CaM-M, and PKA, but not CaMKII, in Ca2+ release termination. Moreover, we showed that isoproterenol and CaM-M suppressed pacing-induced Ca2+ alternans and accelerated Ca2+ transient recovery in intact working hearts, whereas CaM-wild-type exerted an opposite effect. The impact of isoproterenol was partially and fully reversed by the PKA inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline-sulfonamide and the CaMKII inhibitor N-[2-[N-(4-chlorocinnamyl)-N-methylaminomethyl]phenyl]-N-(2-hydroxyethyl)-4-methoxybenzenesulfonamide individually and together, respectively. S2030L abolished the impact of CaM-wild-type, CaM-M, and N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline-sulfonamide-sensitive component, but not the N-[2-[N-(4-chlorocinnamyl)-N-methylaminomethyl]phenyl]-N-(2-hydroxyethyl)-4-methoxybenzenesulfonamide-sensitive component, of isoproterenol.
Filiaciones:
Wei, JH:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Northwest Univ, Sch Med, Xian, Peoples R China
Northwest Univ, Sch Med, Xian 710069, Peoples R China
Guo, WT:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Wang, RW:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Estillore, JP:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Belke, D:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Chen, YX:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Vallmitjana, A:
Univ Politecn Cataluna, Dept Automat Control, Barcelona 08034, Spain
Benitez, R:
Univ Politecn Cataluna, Dept Automat Control, Barcelona 08034, Spain
Hove-Madsen, L:
Hosp Santa Creu & Sant Pau, Biomed Res Inst Barcelona IIBB, CSIC, IIB St Pau, Barcelona 08025, Spain
Hosp Santa Creu & Sant Pau, CIBERCV, Barcelona 08025, Spain
Chen, SRW:
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, Calgary, AB T2N 4N1, Canada
Univ Calgary, Libin Cardiovasc Inst, Dept Physiol & Pharmacol, 3330 Hosp Dr NW, Calgary, AB T2N 4N1, Canada
Bronze, Green Submitted, All Open Access; Bronze Open Access; Green Open Access
|