Phospholamban (PLN), the reversible inhibitor of the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2a), is a key regulator of myocyte Ca(2+) cycling with a significant role in heart failure. We previously showed that the single amino acid difference between human and mouse PLN results in increased inhibition of Ca(2+) cycling and cardiac remodeling and attenuated stress responses in transgenic mice expressing the human PLN (hPLN) in the null background. Here we dissect the molecular and electrophysiological processes triggered by the superinhibitory hPLN in the mouse. Using a multidisciplinary approach, we performed global gene expression analysis, electrophysiology, and mathematical simulations on hPLN mice. We identified significant changes in a series of Na(+) and K(+) homeostasis genes/proteins (including Kcnd2, Scn9a, Slc8a1) and ionic conductance (including L-type Ca(2+) current, Na(+)/Ca(2+) exchanger, transient outward K(+) current). Simulation analysis suggests that this electrical remodeling has a critical role in rescuing cardiac function by improving sarcoplasmic reticulum Ca(2+) load and overall Ca(2+) dynamics. Furthermore, multiple structural and transcription factor gene expression changes indicate an ongoing structural remodeling process, favoring hypertrophy and myogenesis while suppressing apoptosis and progression to heart failure. Our findings expand current understanding of the hPLN function and provide additional insights into the downstream implications of SERCA2a superinhibition in the mammalian heart.
The discovery of potential microarray markers, which will expedite molecular diagnosis/prognosis and provide reliable results to clinical decision-making and treatment selection for patients, is of paramount importance. Feature selection techniques, which aim at minimizing the dimensionality of the microarray data by keeping the most statistically significant genes, are a powerful approach toward this goal. In this paper, we investigate the minimum required subsets of genes, which best classify neuromuscular disease data. For this purpose, we implemented a methodology pipeline that facilitated the use of multiple feature selection methods and subsequent performance of data classification. Five feature selection methods on datasets from ten different neuromuscular diseases were utilized. Our findings reveal subsets of very small number of genes, which can successfully classify normal/disease samples. Interestingly, we observe that similar classification results may be obtained from different subsets of genes. The proposed methodology can expedite the identification of small gene subsets with high-classification accuracy that could ultimately be used in the genetics clinics for diagnostic, prognostic, and pharmacogenomic purposes.
BACKGROUND: Heat shock proteins (Hsp) are known to enhance cell survival under various stress conditions. In the heart, the small Hsp20 has emerged as a key mediator of protection against apoptosis, remodeling, and ischemia/reperfusion injury. Moreover, Hsp20 has been implicated in modulation of cardiac contractility ex vivo. The objective of this study was to determine the in vivo role of Hsp20 in the heart and the mechanisms underlying its regulatory effects in calcium (Ca) cycling. METHODS AND RESULTS: Hsp20 overexpression in intact animals resulted in significant enhancement of cardiac function, coupled with augmented Ca cycling and sarcoplasmic reticulum Ca load in isolated cardiomyocytes. This was associated with specific increases in phosphorylation of phospholamban (PLN) at both Ser16 and Thr17, relieving its inhibition of the apparent Ca affinity of SERCA2a. Accordingly, the inotropic effects of Hsp20 were abrogated in cardiomyocytes expressing nonphosphorylatable PLN (S16A/T17A). Interestingly, the activity of type 1 protein phosphatase (PP1), a known regulator of PLN signaling, was significantly reduced by Hsp20 overexpression, suggesting that the Hsp20 stimulatory effects are partially mediated through the PP1-PLN axis. This hypothesis was supported by cell fractionation, coimmunoprecipitation, and coimmunolocalization studies, which revealed an association between Hsp20, PP1, and PLN. Furthermore, recombinant protein studies confirmed a physical interaction between AA 73 to 160 in Hsp20 and AA 163 to 330 in PP1. CONCLUSIONS: Hsp20 is a novel regulator of sarcoplasmic reticulum Ca cycling by targeting the PP1-PLN axis. These findings, coupled with the well-recognized cardioprotective role of Hsp20, suggest a dual benefit of targeting Hsp20 in heart disease.
The histidine-rich calcium binding protein (HRC) is a novel regulator of sarcoplasmic reticulum (SR) Ca(2+)-uptake, storage and release. Residing in the SR lumen, HRC binds Ca(2+) with high capacity but low affinity. In vitro phosphorylation of HRC affects ryanodine affinity of the ryanodine receptor (RyR), suggesting a functional role of HRC on SR Ca(2+)-release. Indeed, acute HRC overexpression in isolated rodent cardiomyocytes decreases Ca(2+)-induced Ca(2+)-release, increases SR Ca(2+)-load, and impairs contractility. The HRC effects on RyR may be regulated by the Ca(2+)-sensitivity of its interaction with triadin. However, HRC also affects the SR Ca(2+)-ATPase, as shown by HRC overexpression in transgenic mouse hearts, which resulted in reduced SR Ca(2+)-uptake rates, cardiac remodeling and hypertrophy. In fact, in vitro generated evidence suggests that HRC directly interacts with SR Ca(2+)-ATPase2, supporting a dual role of HRC in Ca(2+)-homeostasis: regulation of both SR Ca(2+)-uptake and Ca(2+)-release. Furthermore, HRC plays an important role in myocyte differentiation and in antiapoptotic cardioprotection against ischemia/reperfusion induced cardiac injury. Interestingly, HRC has been linked with familiar cardiac conduction disease and an HRC polymorphism was shown to associate with malignant ventricular arrhythmias in the background of idiopathic dilated cardiomyopathy. This review summarizes studies, which have established the critical role of HRC in Ca(2+)-homeostasis, suggesting its importance in cardiac physiology and pathophysiology.
mailing address: Medical School National and Kapodistrian University of Athens 75 Mikras Asias (Building 15) Athens, 115-27 Greece