Zusammenfassung
Mutations in human cardiac calsequestrin (CASQ2), a high-capacity
calcium-binding protein located in the sarcoplasmic reticulum (SR),
have recently been linked to effort-induced ventricular arrhythmia
and sudden death (catecholaminergic polymorphic ventricular tachycardia).
However, the precise mechanisms through which these mutations affect
SR function and lead to arrhythmia are presently unknown. In this
study, we explored the effect of adenoviral-directed expression of
a canine CASQ2 protein carrying the catecholaminergic polymorphic
ventricular tachycardia-linked mutation D307H (CASQ2(D307H)) on Ca$^2+$
signaling in adult rat myocytes. Total CASQ2 protein levels were
consistently elevated approximately 4-fold in cells infected with
adenoviruses expressing either wild-type CASQ2 (CASQ2(WT)) or CASQ2(D307H).
Expression of CASQ2(D307H) reduced the Ca$^2+$ storing capacity
of the SR. In addition, the amplitude, duration, and rise time of
macroscopic I(Ca)-induced Ca$^2+$ transients and of spontaneous
Ca$^2+$ sparks were reduced significantly in myocytes expressing
CASQ2(D307H). Myocytes expressing CASQ2(D307H) also displayed drastic
disturbances of rhythmic oscillations in Ca$^2+$i and membrane
potential, with signs of delayed afterdepolarizations when undergoing
periodic pacing and exposed to isoproterenol. Importantly, normal
rhythmic activity was restored by loading the SR with the low-affinity
Ca$^2+$ buffer, citrate. Our data suggest that the arrhythmogenic
CASQ2(D307H) mutation impairs SR Ca$^2+$ storing and release
functions and destabilizes the Ca$^2+$-induced Ca$^2+$ release
mechanism by reducing the effective Ca$^2+$ buffering inside
the SR and/or by altering the responsiveness of the Ca$^2+$ release
channel complex to luminal Ca$^2+$. These results establish at
the cellular level the pathological link between CASQ2 mutations
and the predisposition to adrenergically mediated arrhythmias observed
in patients carrying CASQ2 defects.
- acid
- adenoviridae,
- amino
- animals;
- calcium
- calsequestrin,
- cardiac,
- cardiac;
- channel
- channel,
- death,
- dogs;
- fusion
- gating;
- genetic
- genetics/metabolism
- genetics/physiology;
- genetics;
- humans;
- ion
- macromolecular
- male;
- metabolism;
- missense;
- mutation,
- mutation;
- myocytes,
- physiology;
- point
- proteins,
- rats,
- rats;
- receptor
- recombinant
- release
- reticulum,
- ryanodine
- sarcoplasmic
- signaling,
- sprague-dawley;
- substances;
- substitution;
- sudden,
- tachycardia,
- vectors,
- ventricular,
Nutzer