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CaMKII-dependent phosphorylation of cardiac ryanodine receptors regulates cell death in cardiac ischemia/reperfusion injury.

ABSTRACT: Ca(2+)-calmodulin kinase II (CaMKII) activation is deleterious in cardiac ischemia/reperfusion (I/R). Moreover, inhibition of CaMKII-dependent phosphorylations at the sarcoplasmic reticulum (SR) prevents CaMKII-induced I/R damage. However, the downstream targets of CaMKII at the SR level, responsible for this detrimental effect, remain unclear. In the present study we aimed to dissect the role of the two main substrates of CaMKII at the SR level, phospholamban (PLN) and ryanodine receptors (RyR2), in CaMKII-dependent I/R injury. In mouse hearts subjected to global I/R (45/120min), phosphorylation of the primary CaMKII sites, S2814 on cardiac RyR2 and of T17 on PLN, significantly increased at the onset of reperfusion whereas PKA-dependent phosphorylation of RyR2 and PLN did not change. Similar results were obtained in vivo, in mice subjected to regional myocardial I/R (1/24h). Knock-in mice with an inactivated serine 2814 phosphorylation site on RyR2 (S2814A) significantly improved post-ischemic mechanical recovery, reduced infarct size and decreased apoptosis. Conversely, knock-in mice, in which CaMKII site of RyR2 is constitutively activated (S2814D), significantly increased infarct size and exacerbated apoptosis. In S2814A and S2814D mice subjected to regional myocardial ischemia, infarct size was also decreased and increased respectively. Transgenic mice with double-mutant non-phosphorylatable PLN (S16A/T17A) in the PLN knockout background (PLNDM) also showed significantly increased post-ischemic cardiac damage. This effect cannot be attributed to PKA-dependent PLN phosphorylation and was not due to the enhanced L-type Ca(2+) current, present in these mice. Our results reveal a major role for the phosphorylation of S2814 site on RyR2 in CaMKII-dependent I/R cardiac damage. In contrast, they showed that CaMKII-dependent increase in PLN phosphorylation during reperfusion opposes rather than contributes to I/R damage.

SUBMITTER: Di Carlo MN

PROVIDER: S-EPMC4131282 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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CaMKII-dependent phosphorylation of cardiac ryanodine receptors regulates cell death in cardiac ischemia/reperfusion injury.

Di Carlo Mariano N MN Said Matilde M Ling Haiyun H Valverde Carlos A CA De Giusti Verónica C VC Sommese Leandro L Palomeque Julieta J Aiello Ernesto A EA Skapura Darlene G DG Rinaldi Gustavo G Respress Jonathan L JL Brown Joan Heller JH Wehrens Xander H T XH Salas Margarita A MA Mattiazzi Alicia A

Journal of molecular and cellular cardiology 20140617

Ca(2+)-calmodulin kinase II (CaMKII) activation is deleterious in cardiac ischemia/reperfusion (I/R). Moreover, inhibition of CaMKII-dependent phosphorylations at the sarcoplasmic reticulum (SR) prevents CaMKII-induced I/R damage. However, the downstream targets of CaMKII at the SR level, responsible for this detrimental effect, remain unclear. In the present study we aimed to dissect the role of the two main substrates of CaMKII at the SR level, phospholamban (PLN) and ryanodine receptors (RyR2 ...[more]

PMID: 24949568

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Project description:RationaleThe levels of a small heat shock protein (Hsp)20 and its phosphorylation are increased on ischemic insults, and overexpression of Hsp20 protects the heart against ischemia/reperfusion injury. However, the mechanism underlying cardioprotection of Hsp20 and especially the role of its phosphorylation in regulating ischemia/reperfusion-induced autophagy, apoptosis, and necrosis remain to be clarified.ObjectiveHerein, we generated a cardiac-specific overexpression model, carrying nonphosphorylatable Hsp20, where serine 16 was substituted with alanine (Hsp20(S16A)). By subjecting this model to ischemia/reperfusion, we addressed whether: (1) the cardioprotective effects of Hsp20 are associated with serine 16 phosphorylation; (2) blockade of Hsp20 phosphorylation influences the balance between autophagy and cell death; and (3) the aggregation pattern of Hsp20 is altered by its phosphorylation.Methods and resultsOur results demonstrated that Hsp20(S16A) hearts were more sensitive to ischemia/reperfusion injury, evidenced by lower recovery of contractile function and increased necrosis and apoptosis, compared with non-TG hearts. Interestingly, autophagy was activated in non-TG hearts but significantly inhibited in Hsp20(S16A) hearts following ischemia/reperfusion. Accordingly, pretreatment of Hsp20(S16A) hearts with rapamycin, an activator of autophagy, resulted in improvement of functional recovery, compared with saline-treated Hsp20(S16A) hearts. Furthermore, on ischemia/reperfusion, the oligomerization pattern of Hsp20 appeared to shift to higher aggregates in Hsp20(S16A) hearts.ConclusionsCollectively, these data indicate that blockade of Ser16-Hsp20 phosphorylation attenuates the cardioprotective effects of Hsp20 against ischemia/reperfusion injury, which may be attributable to suppressed autophagy and increased cell death. Therefore, phosphorylation of Hsp20 at serine 16 may represent a potential therapeutic target in ischemic heart disease.

Dataset Information

CaMKII-dependent phosphorylation of cardiac ryanodine receptors regulates cell death in cardiac ischemia/reperfusion injury.

Publications

CaMKII-dependent phosphorylation of cardiac ryanodine receptors regulates cell death in cardiac ischemia/reperfusion injury.

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