Regulation of Ca2+ signaling in transgenic mouse cardiac myocytes overexpressing calsequestrin.

Abstract

To probe the physiological role of calsequestrin in excitation-contraction coupling, transgenic mice overexpressing cardiac calsequestrin were developed. Transgenic mice exhibited 10-fold higher levels of calsequestrin in myocardium and survived into adulthood, but had severe cardiac hypertrophy, with a twofold increase in heart mass and cell size. In whole cell-clamped transgenic myocytes, Ca2+ channel- gated Ca2+ release from the sarcoplasmic reticulum was strongly suppressed, the frequency of occurrence of spontaneous or Ca2+ current-triggered "Ca2+ sparks" was reduced, and the spark perimeter was less defined. In sharp contrast, caffeine-induced Ca2+ transients and the resultant Na+-Ca2+ exchanger currents were increased 10-fold in transgenic myocytes, directly implicating calsequestrin as the source of the contractile-dependent pool of Ca2+. Interestingly, the proteins involved in the Ca2+-release cascade (ryanodine receptor, junctin, and triadin) were downregulated, whereas Ca2+-uptake proteins (Ca2+-ATPase and phospholamban) were unchanged or slightly increased. The parallel increase in the pool of releasable Ca2+ with overexpression of calsequestrin and subsequent impairment of physiological Ca2+ release mechanism show for the first time that calsequestrin is both a storage and a regulatory protein in the cardiac muscle Ca2+-signaling cascade. Cardiac hypertrophy in these mice may provide a novel model to investigate the molecular determinants of heart failure.

Ca2+ signalling and muscle disease

“…Loss of SERCA1a Ca 21 pump function is one cause of exercise-induced impairment of the relaxation of skeletal muscle, the major diagnostic feature of Brody disease [17]. Studies with knockout and transgenic mice show that gain of inhibitory function of phospholamban, a regulator of the Ca 21 pump, alters cardiac contractility [18,19] and could be a causal feature in some cardiomyopathies, that overexpression of cardiac calsequestrin leads to cardiomyopathy [20] and that ablation of calreticulin alters cardiac development [21]. Darier's disease [OMIM #124200] [18±19] is an autosomal dominant skin disorder, characterized by a loss of adhesion between epithelial cells that is associated with breakdown of desmosomal filament complexes and premature keratinization.…”

“…Calsequestrin is clearly designed to be a high capacity, low affinity Ca 21 binding protein [12]. Transgenic mice exhibiting 10-fold overexpression of cardiac calsequestrin developed severe cardiac hypertrophy, with a twofold increase in heart mass and cell size [20]. In whole cell-clamped transgenic myocytes, Ca 21 channel-gated Ca 21 release through the ryanodine receptor was strongly suppressed and the frequency of`Ca 21 sparks' was reduced.…”

Increased Ca2+ storage capacity of the skeletal muscle sarcoplasmic reticulum of transgenic mice over‐expressing membrane bound calcium binding protein junctate

“…These findings are consistent with other investigations in which the effect of over-expression of SR/ER high capacity low affinity Ca 2þ binding properties was studied. Indeed chronic over-expression of calsequestrin, the major Ca 2þ binding protein in striated muscle in cardiomyocytes, increased the Ca 2þ storage capacity of the SR but did not affect the expression level of SERCA or phospholamban (Jones et al, 1998;Sato et al, 1998;Knollmann et al, 2000;Miller et al, 2005). Consistent results showing an increase in the lumenal SR Ca 2þ buffering capacity were obtained when the luminal histidine-rich Ca 2þ binding protein (HRC) was over-expressed in neonatal and adult rat cardiomyocytes (Kim et al, 2003;Fan et al, 2004).…”

“…Furthermore, over-expression of other known calcium storage proteins such as calsequestrin and histidine rich Ca 2þ binding protein leads to an increase in caffeine-induced Ca 2þ release (Jones et al, 1998;Sato et al, 1998;Kim et al, 2003;Fan et al, 2004;Miller et al, 2005). Over-expression of junctate also occurs in a membrane compartment endowed with the RyR Ca 2þ release channel, since we found that (i) direct stimulation of isolated myotubes with either caffeine or 4-CmC resulted in a significantly larger Ca 2þ transient and (ii) total SR microsomes from transgenic mice show a 24% increase of the extent of calcium release induced by 4-CmC.…”

Calcium cycling proteins in heart failure, cardiomyopathy and arrhythmias

“…To elucidate the physiological significance of cardiac CSQ (CSQ2) in the cardiac excitation-contraction coupling, two independent lines of transgenic mice overexpressing CSQ2 in the heart have been generated (Jones et al, 1998;Sato et al, 1998). Although cardiac-targeted overexpression of CSQ2 results in a marked increase in SR Ca 2+ storage in the both models, SR Ca 2+ release was impaired upon depolarization, leading to depressed contractile parameters and cardiac hypertrophy.…”