Cardiac myocyte Z-line calmodulin is mainly RyR2-bound, and reduction is arrhythmogenic and occurs in heart failure

Yi Yang, Tao Guo, Tetsuro Oda, Asima Chakraborty, Le Chen, Hitoshi Uchinoumi, Anne A. Knowlton, Bradley R. Fruen, Razvan L. Cornea, Gerhard Meissner, Donald M. Bers

Research output: Contribution to journalArticlepeer-review

49 Scopus citations

Abstract

Rationale: Calmodulin (CaM) associates with cardiac ryanodine receptor type-2 (RyR2) as an important regulator. Defective CaM-RyR2 interaction may occur in heart failure, cardiac hypertrophy, and catecholaminergic polymorphic ventricular tachycardia. However, the in situ binding properties for CaM-RyR2 are unknown. Objective: We sought to measure the in situ binding affinity and kinetics for CaM-RyR2 in normal and heart failure ventricular myocytes, estimate the percentage of Z-line-localized CaM that is RyR2-bound, and test cellular function of defective CaM-RyR2 interaction. Methods and Results: Using fluorescence resonance energy transfer in permeabilized myocytes, we specifically resolved RyR2-bound CaM from other potential binding targets and measured CaM-RyR2 binding affinity in situ (Kd=10-20 nmol/L). Using RyR2ADA/+ knock-in mice, in which half of the CaM-RyR2 binding is suppressed, we estimated that >90% of Z-line CaM is RyR2-bound. Functional tests indicated a higher propensity for Ca2+ wave production and stress-induced ventricular arrhythmia in RyRADA/+ mice. In a post-myocardial infarction rat heart failure model, we detected a decrease in the CaM-RyR2 binding affinity (Kd≈51 nmol/L; ≈3-fold increase) and unaltered RyR2 affinity for the FK506-binding protein FKBP12.6 (Kd0.8 nmol/L). Conclusions: CaM binds to RyR2 with high affinity in cardiac myocytes. Physiologically, CaM is bound to >70% of RyR2 monomers and inhibits sarcoplasmic reticulum Ca2+ release. RyR2 is the major binding site for CaM along the Z-line in cardiomyocytes, and dissociating CaM from RyR2 can cause severe ventricular arrhythmia. In heart failure, RyR2 shows decreased CaM affinity, but unaltered FKBP 12.6 affinity.

Original languageEnglish (US)
Pages (from-to)295-306
Number of pages12
JournalCirculation research
Volume114
Issue number2
DOIs
StatePublished - Jan 1 2014

Keywords

  • Arrhythmias
  • Cardiac
  • Fluorescence resonance energy transfer
  • Heart failure
  • Ryanodine receptor calcium release channel

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