RESUMEN
S100A1 is an interesting Ca2+ binding protein with respect to muscle physiology as it is preferentially expressed in cardiac muscle and colocalizes with the sarcolemmal and the sarcoplasmic reticulum membranes as well as with the sarcomere. It is therefore conceivable that S100A1 may play a specific role in the regulation of cardiac Ca2+ homeostasis and contractility. We therefore investigated the impact of adenoviral S100A1 overexpression on fractional shortening (FS%) and systolic Ca2+ transients in adult rat cardiomyocytes as well as of S100A1 protein on SERCA activity in skinned cell preparation. In our setting S100A1 gene transfer increased FS% by 55%, systolic Ca2+ amplitudes by 62%, while S100A1 protein increased SERCA activity by 28%. Importantly, the gain in systolic Ca2+ supply was not only seen on basal conditions but also with isoproterenol-stimulated Ca2+ cycling. Thus, S100A1 enhances cardiac contractility by increasing intracellular Ca2+ fluxes at least in part due to a modulation of SERCA. Since earlier observations demonstrated S100A1 protein levels to be increased in compensatory hypertrophy and significantly downregulated in end stage heart failure, these functional data suggest that S100A1 is a novel determinant of cardiac function whose expression levels are causally related to the prevailing contractile state of the heart.
Asunto(s)
Proteínas de Unión al Calcio/farmacología , Calcio/metabolismo , Contracción Miocárdica/efectos de los fármacos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/fisiología , Adenoviridae/genética , Animales , Proteínas de Unión al Calcio/genética , Células Cultivadas , Vectores Genéticos , Homeostasis/efectos de los fármacos , Humanos , Ratas , Proteínas S100 , Retículo Sarcoplasmático/metabolismo , TransfecciónRESUMEN
S100A1, a Ca2+-binding protein of the EF-hand type, is most highly expressed in striated muscle and has previously been shown to interact with the skeletal muscle sarcoplasmic reticulum (SR) Ca2+ release channel/ryanodine receptor (RyR1) isoform. However, it was unclear whether S100A1/RyR1 interaction could modulate SR Ca2+ handling and contractile properties in skeletal muscle fibers. Since S100A1 protein is differentially expressed in fast- and slow-twitch skeletal muscle, we used saponin-skinned murine Musculus extensor digitorum longus (EDL) and Musculus soleus (Soleus) fibers to assess the impact of S100A1 protein on SR Ca2+ release and isometric twitch force in functionally intact permeabilized muscle fibers. S100A1 equally enhanced caffeine-induced SR Ca2+ release and Ca2+-induced isometric force transients in both muscle preparations in a dose-dependent manner. Introducing a synthetic S100A1 peptide model (devoid of EF-hand Ca2+-binding sites) allowed identification of the S100A1 C terminus (amino acids 75-94) and hinge region (amino acids 42-54) to differentially enhance SR Ca2+ release with a nearly 3-fold higher activity of the C terminus. These effects were exclusively based on enhanced SR Ca2+ release as S100A1 influenced neither SR Ca2+ uptake nor myofilament Ca2+ sensitivity/cooperativity in our experimental setting. In conclusion, our study shows for the first time that S100A1 augments contractile performance both of fast- and slow-twitch skeletal muscle fibers based on enhanced SR Ca2+ efflux at least mediated by the C terminus of S100A1 protein. Thus, our data suggest that S100A1 may serve as an endogenous enhancer of SR Ca2+ release and might therefore be of physiological relevance in the process of excitation-contraction coupling in skeletal muscle.
Asunto(s)
Proteínas de Unión al Calcio/química , Proteínas de Unión al Calcio/metabolismo , Músculo Esquelético/metabolismo , Secuencia de Aminoácidos , Animales , Cafeína/farmacología , Señalización del Calcio/efectos de los fármacos , Proteínas de Unión al Calcio/genética , Humanos , Técnicas In Vitro , Contracción Isométrica/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Fibras Musculares de Contracción Rápida/efectos de los fármacos , Fibras Musculares de Contracción Rápida/metabolismo , Fibras Musculares de Contracción Lenta/efectos de los fármacos , Fibras Musculares de Contracción Lenta/metabolismo , Músculo Esquelético/efectos de los fármacos , Fragmentos de Péptidos/química , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Proteínas S100 , Retículo Sarcoplasmático/efectos de los fármacos , Retículo Sarcoplasmático/metabolismoRESUMEN
S100A1, a Ca2+-sensing protein of the EF-hand family, is most highly expressed in myocardial tissue, and cardiac S100A1 overexpression in vitro has been shown to enhance myocyte contractile properties. To study the physiological consequences of S100A1 in vivo, transgenic mice were developed with cardiac-restricted overexpression of S100A1. Characterization of two independent transgenic mouse lines with approximately 4-fold overexpression of S100A1 in the myocardium revealed a marked augmentation of in vivo basal cardiac function that remained elevated after beta-adrenergic receptor stimulation. Contractile function and Ca2+ handling properties were increased in ventricular cardiomyocytes isolated from S100A1 transgenic mice. Enhanced cellular Ca2+ cycling by S100A1 was associated both with increased sarcoplasmic reticulum Ca2+ content and enhanced sarcoplasmic reticulum Ca2+-induced Ca2+ release, and S100A1 was shown to associate with the cardiac ryanodine receptor. No alterations in beta-adrenergic signal transduction or major cardiac Ca2+-cycling proteins occurred, and there were no signs of hypertrophy with chronic cardiac S100A1 overexpression. Our findings suggest that S100A1 plays an important in vivo role in the regulation of cardiac function perhaps through interacting with the ryanodine receptor. Because S100A1 protein expression is down-regulated in heart failure, increasing S100A1 expression in the heart may represent a novel means to augment contractility.