Early cardiac hypertrophy in mice with impaired calmodulin regulation of cardiac muscle Ca2+ release channel
Naohiro Yamaguchi, … , Oliver Smithies, Gerhard Meissner
Naohiro Yamaguchi, … , Oliver Smithies, Gerhard Meissner
Published May 1, 2007
Citation Information: J Clin Invest. 2007;117(5):1344-1353. https://doi.org/10.1172/JCI29515.
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Research Article Cardiology

Early cardiac hypertrophy in mice with impaired calmodulin regulation of cardiac muscle Ca2+ release channel

Abstract

Studies with isolated membrane fractions have shown that calmodulin (CaM) inhibits the activity of cardiac muscle cell Ca2+ release channel ryanodine receptor 2 (RyR2). To determine the physiological importance of CaM regulation of RyR2, we generated a mouse with 3 amino acid substitutions (RyR2-W3587A/L3591D/F3603A) in exon 75 of the Ryr2 gene, which encodes the CaM-binding site of RyR2. Homozygous mutant mice showed an increased ratio of heart weight to body weight, greatly reduced fractional shortening of the left ventricle, and lethality at 9–16 days of age. Biochemical analysis of hearts of 7- and 10-day-old homozygous mutant mice indicated an impaired CaM inhibition of RyR2 at micromolar Ca2+ concentrations, reduction in RyR2 protein levels and sarcoplasmic reticulum Ca2+ sequestration, and upregulation of genes and/or proteins associated with class II histone deacetylase/myocyte enhancer factor-2 and calcineurin signaling pathways. Sustained Ca2+ transients, often displaying repeated periods of incomplete Ca2+ removal, were observed in homozygous cardiomyocytes. Taken together, the data indicate that impaired CaM inhibition of RyR2, associated with defective sarcoplasmic reticulum Ca2+ release and altered gene expression, leads to cardiac hypertrophy and early death.

Authors

Naohiro Yamaguchi, Nobuyuki Takahashi, Le Xu, Oliver Smithies, Gerhard Meissner

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Figure 5

CaM binding and regulation of RyR2 from hearts of WT and mutant mice.

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CaM binding and regulation of RyR2 from hearts of WT and mutant mice. (A...
(A) Western blots of RyR2 bound to CaM-Sepharose beads. Input RyR2s (left) and RyR2s bound to CaM-Sepharose in the absence (–) or presence (+) of 2 μM CaM (right). (B and C) Specific [3H]ryanodine binding to crude membrane fractions from hearts of 7-day-old and 10-day-old WT, heterozygous, and homozygous mice was determined as described in Supplemental Methods. (B) Ca2+ dependence of [3H]ryanodine binding in absence of CaM. (C) [3H]ryanodine binding as a function of CaM concentration at 1.2 μM Ca2+. Data are the mean ± SEM of 4–6 experiments. *P < 0.05, #P < 0.01 compared with control (no CaM added), as determined by Student’s t test.