PDIA3的结构与功能及其在精卵融合过程中的作用

二硫键异构酶家族的一员PDIA3(protein disulfide-isomerase A3)最初确定其位于内质网腔内,作为钙联接蛋白和钙网质蛋白的分子伴侣复合物的组成成分之一,促进新合成的糖蛋白在内质网中的正确折叠。PDIA3在内质网中的功能依赖其蛋白二硫键氧化还原异构酶活性。目前研究发现位于精子表面的PDIA3的蛋白二硫键氧化还原酶活性在精卵融合中也发挥着重要的作用。就目前关于PDIA3的结构和功能及在精卵质膜融合中的作用研究进展做一概述。     

Stress-induced expression of protein disulfide isomerase associated 3 (PDIA3) in Atlantic salmon (Salmo salar L.)

In mammals, disulfide isomerase associated 3, PDIA3, is a member of the endoplasmic reticulum (ER) stress proteins, which can be induced by oxidative stress; however, its role in relation to stress regulation is still unknown in fish. Here, we report the cloning of a coding region of PDIA3 from the Atlantic salmon. PDIA3 mRNA expression was evaluated in the liver of Atlantic salmon exposed to environmental hyperoxia stress and toxic perfluorooctane sulfonate (PFOS) exposure stress. The PDIA3 sequence contained two PDI-typical thioredoxin active sites of WCGHC and shared approximately 70% identity with mammalian PDIA3, and its mRNA was primarily expressed in the liver. PDIA3 was significantly increased in the liver of Atlantic salmon exposed to hyperoxic water during smoltification. Also Mn superoxide dismutase (Mn-SOD) and CCAAT/enhancer binding protein (C/EBP), other markers of oxidative stress, were upregulated by hyperoxia. Furthermore, PFOS exposure of hepatocytes resulted in elevated mRNA expression of PDIA3, Mn-SOD and C/EBPδ as well as peroxisome proliferator-activated receptor gamma (PPARγ). These results indicate a signaling connection between oxidative stress and ER stress. PDIA3 and C/EBPδ may be valuable markers in fish for exposure and effect to environmental stress.     

Overexpression of C-terminal Src kinase blocks 14, 15-epoxyeicosatrienoic acid-induced tyrosine phosphorylation and mitogenesis

We have previously reported that 14,15-epoxyeicosatrienoic acid (14, 15-EET) is a potent mitogen for the renal epithelial cell line, LLCPKcl4. This mitogenic effect is dependent upon activation of a protein-tyrosine kinase cascade that results in activation of mitogen-activated protein kinase and phosphatidylinositol 3-kinase. Because of suggestive evidence that 14,15-EET also activated Src in these cells, we stably transfected LLCPKcl4 with an expression construct of the C-terminal Src kinase (CSK), which inhibits Src family kinase activity. In vitro Src kinase activity assays confirmed that in empty vector-transfected cells (Vector cells), 14, 15-EET increased Src kinase activity, while in clones overexpressing CSK mRNA and immunoreactive protein (CSK cells), 14,15-EET-induced activation of Src was almost completely blocked (94% inhibition). Of interest, epidermal growth factor (EGF) and fetal bovine serum (FBS) also increased Src activity in Vector cells, but not in CSK cells, further confirming the ability of CSK overexpression to prevent Src activation. CSK cells failed to increase [(3)H]thymidine incorporation in response to exogenous 14,15-EET. In contrast, both EGF and FBS significantly increased [(3)H]thymidine incorporation in CSK cells. Immunoprecipitation with anti-phosphotyrosine antibodies and immunoblotting with an antibody against extracellular signal-regulated kinase (ERK) indicated that in CSK cells, 14,15-EET failed to activate ERK1 and ERK2; however, EGF- and FBS-induced activation of ERKs was not different from that seen in Vector cells. In Vector cells, the 14,15-EET-stimulated tyrosine phosphorylation of ERKs was blocked by pretreatment with 1 microm PP2, a selective inhibitor of Src kinases. The present study demonstrates that 14, 15-EET exerts its mitogenic effects predominantly through a Src kinase-mediated pathway, which is the most upstream signaling step determined to date in the 14,15-EET-activated tyrosine kinase cascade in renal epithelial cells.     

Increased myocardial expression of RAMP1 and RAMP3 in rats with chronic heart failure

Calcitonin gene-related peptide (CGRP) and adrenomedullin (ADM) are potent vasodilators in humans and improved myocardial ischemia is observed after CGRP administration. Receptors for CGRP and ADM were already identified in heart. Receptor activity-modifying proteins (RAMPs) determine the ligand specificity of the calcitonin receptor-like receptor (CRLR); co-expression of RAMP1 and CRLR results in a CGRP receptor, whereas the association of RAMP2 or RAMP3 with CRLR gives an ADM receptor. As CGRP and ADM may play a beneficial role in heart failure, we investigated whether the CGRP and ADM receptors are upregulated in chronic heart failure. We have used semi-quantitative RT-PCR and Western-blot analysis to detect and quantify the mRNA and the protein of RAMP1 and RAMP3 in both atria and ventricles of failing hearts 6 months after aortic banding in rats. Our results showed for the first time an up-regulation of RAMP1 and RAMP3 mRNAs and proteins in this model of cardiac failure. No change was observed in mRNAs coding for CRLR, RAMP2, RDC1 (canine orphan receptor), and ADM. The present results suggested after congestive heart failure in adult rats, an up-regulation of the CGRP receptor (by an increase in RAMP1 that is associated with CRLR) in atria and ventricles and of ADM receptor (by increased RAMP3 expression that is associated with CRLR) in atria. These findings support a functional role for CGRP and ADM receptors to compensate the chronic heart failure in rats.     

Upregulation of beta-adrenergic receptors in heart failure due to volume overload

To examine the mechanisms of changes in beta-adrenergic signal transduction in heart failing due to volume overload, we studied the status of beta-adrenoceptors (beta-ARs), G protein-coupled receptor kinase (GRK), and beta-arrestin in heart failure due to aortocaval shunt (AVS). Heart failure in rats was induced by creating AVS for 16 wk, and beta-AR binding, GRK activity, as well as their protein content, and mRNA levels were determined in both left and right ventricles. The density and protein content for beta1-ARs, unlike those for beta2-ARs, were increased in the failing hearts. Furthermore, protein contents for GRK isoforms and beta-arrestin-1 were decreased in membranous fractions and increased in cytosolic fractions from the failing hearts. On the other hand, steady-state mRNA levels for beta1-ARs and GRK2, as well as protein content for Gbetagamma-subunits, did not change in the failing heart. Basal cardiac function was depressed; however, both in vivo and ex vivo positive inotropic responses of the failing hearts to isoproterenol were augmented. Treatment of AVS animals with imidapril (1 mg.kg(-1).day(-1)) or losartan (20 mg.kg(-1).day(-1)) retarded the progression of heart failure; partially prevented changes in beta1-ARs, GRKs, and beta-arrestin-1 in the failing myocardium; and attenuated the increase in positive inotropic effect of isoproterenol. These results indicate that upregulation of beta1-ARs is associated with subcellular redistribution of GRKs and beta-arrestin-1 in the failing heart due to volume overload. Furthermore, attenuation of alterations in beta-adrenergic system by imidapril or losartan may be due to blockade of the renin-angiotensin system in the AVS model of heart failure.     

Role of Ca2+ in diperoxovanadate-induced cytoskeletal remodeling and endothelial cell barrier function

Diperoxovanadate (DPV), a potent inhibitor of protein tyrosine phosphatases and activator of tyrosine kinases, alters endothelial barrier function via signaling pathways that are incompletely understood. One potential pathway is Src kinase-mediated tyrosine phosphorylation of proteins such as cortactin that regulate endothelial cell (EC) cytoskeleton assembly. As DPV modulates endothelial cell signaling via protein tyrosine phosphorylation, we determined the role of DPV-induced intracellular free calcium concentration ([Ca2+]i) in activation of Src kinase, cytoskeletal remodeling, and barrier function in bovine pulmonary artery endothelial cells (BPAECs). DPV in a dose- and time-dependent fashion increased [Ca2+]i, which was partially blocked by the calcium channel blockers nifedipine and Gd3+. Treatment of cells with thapsigargin released Ca2+ from the endoplasmic reticulum, and subsequent addition of DPV caused no further change in [Ca2+]i. These data suggest that DPV-induced [Ca2+]i includes Ca release from the endoplasmic reticulum and Ca influx through store-operated calcium entry. Furthermore, DPV induced an increase in protein tyrosine phosphorylation, phosphorylation of Src and cortactin, actin remodeling, and altered transendothelial electrical resistance in BPAECs. These DPV-mediated effects were significantly attenuated by BAPTA (25 microM), a chelator of [Ca2+]i. Immunofluorescence studies reveal that the DPV-mediated colocalization of cortactin with peripheral actin was also prevented by BAPTA. Chelation of extracellular Ca2+ by EGTA had marginal effects on DPV-induced phosphorylation of Src and cortactin; actin stress fibers formation, however, affected EC barrier function. These data suggest that DPV-induced changes in [Ca2+]i regulate endothelial barrier function using signaling pathways that involve Src and cytoskeleton remodeling.     

Molecular correlates of altered expression of potassium currents in failing rabbit myocardium

Action potential (AP) prolongation is a hallmark of failing myocardium. Functional downregulation of K currents is a prominent feature of cells isolated from failing ventricles. The detailed changes in K current expression differ depending on the species, the region of the heart, and the mechanism of induction of heart failure. We used complementary approaches to study K current downregulation in pacing tachycardia-induced heart failure in the rabbit. The AP duration (APD) at 90% repolarization was significantly longer in cells isolated from failing hearts compared with controls (539 +/- 162 failing vs. 394 +/- 114 control, P < 0.05). The major K currents in the rabbit heart, inward rectifier potassium current (I(K1)), transient outward (I(to)), and delayed rectifier current (I(K)) were functionally downregulated in cells isolated from failing ventricles. The mRNA levels of Kv4.2, Kv1.4, KChIP2, and Kir2.1 were significantly downregulated, whereas the Kv4.3, Erg, KvLQT1, and minK were unaltered in the failing ventricles compared with the control left ventricles. Significant downregulation in the long splice variant of Kv4.3, but not in the total Kv4.3, Kv4.2, and KChIP2 immunoreactive protein, was observed in cells isolated from the failing ventricle with no change in Kv1.4, KvLQT1, and in Kir2.1 immunoreactive protein levels. Multiple cellular and molecular mechanisms underlie the downregulation of K currents in the failing rabbit ventricle.     

Integrating cardiac PIP3 and cAMP signaling through a PKA anchoring function of p110γ

Adrenergic stimulation of the heart engages cAMP and phosphoinositide second messenger signaling cascades. Cardiac phosphoinositide 3-kinase p110γ participates in these processes by sustaining β-adrenergic receptor internalization through its catalytic function and by controlling phosphodiesterase 3B (PDE3B) activity via an unknown kinase-independent mechanism. We have discovered that p110γ anchors protein kinase A (PKA) through a site in its N-terminal region. Anchored PKA activates PDE3B to enhance cAMP degradation and phosphorylates p110γ to inhibit PIP(3) production. This provides local feedback control of PIP(3) and cAMP signaling events. In congestive heart failure, p110γ is upregulated and escapes PKA-mediated inhibition, contributing to a reduction in β-adrenergic receptor density. Pharmacological inhibition of p110γ normalizes β-adrenergic receptor density and improves contractility in failing hearts.     

Structure of the Non-Catalytic Domain of the Protein Disulfide Isomerase-Related Protein (PDIR) Reveals Function in Protein Binding

Protein disulfide isomerases comprise a large family of enzymes responsible for catalyzing the proper oxidation and folding of newly synthesized proteins in the endoplasmic reticulum (ER). Protein disulfide isomerase-related (PDIR) protein (also known as PDIA5) is a specialized member that participates in the folding of α₁-antitrypsin and N-linked glycoproteins. Here, the crystal structure of the non-catalytic domain of PDIR was determined to 1.5 Å resolution. The structure adopts a thioredoxin-like fold stabilized by a structural disulfide bridge with a positively charged binding surface for interactions with the ER chaperones, calreticulin and ERp72. Crystal contacts between molecules potentially mimic the interactions of PDIR with misfolded substrate proteins. The results suggest that the non-catalytic domain of PDIR plays a key role in the recognition of protein partners and substrates.     

Thapsigargin down-regulates protein levels of GRP78/BiP in INS-1E cells

Pancreatic β-cells have a well-developed endoplasmic reticulum (ER) and express large amounts of chaperones and protein disulfide isomerases (PDI) to meet the high demand for synthesis of proteins. We have observed an unexpected decrease in chaperone protein level in the β-cell model INS-1E after exposure to the ER stress inducing agent thapsigargin. As these cells are a commonly used model for primary β-cells and has been shown to be vulnerable to ER stress, we hypothesize these cells are incapable of mounting a chaperone defense upon activation of ER stress. To investigate the chaperone expression during an ER stress response, induced by thapsigargin in INS-1E cells, we used quantitative mass spectrometry based proteomics. The results displayed a decrease of GRP78/BiP, PDIA3 and PDIA6. Decrease of GRP78/BiP was verified by Western blot and occurred in parallel with enhanced levels of p-eIF2α and CHOP. In contrast to INS-1E cells, GRP78/BiP was not decreased in MIN6 cell or rat and mouse islets after thapsigargin exposure. Investigation of the decreased protein levels of GRP78/BiP indicates that this is not a consequence of reduced mRNA expression. Rather the reduction results from the combined effect of reduced protein synthesis and enhanced proteosomal degradation and possibly also degradation via autophagy. Induction of ER stress with thapsigargin leads to lower protein levels of GRP78/BiP, PDIA3 and PDIA6 in INS-1E cells which may contribute to the susceptibility of ER stress in this β-cell model.     

Src regulates Golgi structure and KDEL receptor-dependent retrograde transport to the endoplasmic reticulum

The tyrosine kinase Src is present on the Golgi membranes. Its role, however, in the overall function and organization of the Golgi apparatus is unclear. We have found that in a cell line called SYF, which lacks the three ubiquitous Src-like kinases (Src, Yes, and Fyn), the organization of the Golgi apparatus is perturbed. The Golgi apparatus is composed of collapsed stacks and bloated cisternae in these cells. Expression of an activated form of Src relocated the KDEL receptor (KDEL-R) from the Golgi apparatus to the endoplasmic reticulum. Other Golgi-specific marker proteins were not affected under these conditions. Because of the specific effect of Src on the location of KDEL-R, we tested whether protein transport between ER and the Golgi apparatus involves Src. Transport of Pseudomonas exotoxin, which is transported to the ER by binding to the KDEL-R is accelerated by inhibition or genetic ablation of Src. Protein transport from ER to the Golgi apparatus however, is unaffected by Src deletion or inhibition. We propose that Src has an appreciable role in the organization of the Golgi apparatus, which may be linked to its involvement in protein transport from the Golgi apparatus to the endoplasmic reticulum.     

β-AR blockers suppresses ER stress in cardiac hypertrophy and heart failure

BACKGROUND: Long-term β-adrenergic receptor (β-AR) blockade reduces mortality in patients with heart failure. Chronic sympathetic hyperactivity in heart failure causes sustained β-AR activation, and this can deplete Ca(2+) in endoplasmic reticulum (ER) leading to ER stress and subsequent apoptosis. We tested the effect of β-AR blockers on ER stress pathway in experimental model of heart failure. METHODS AND DISCUSSIONS: ER chaperones were markedly increased in failing hearts of patients with end-stage heart failure. In Sprague-Dawley rats, cardiac hypertrophy and heart failure was induced by abdominal aortic constriction or isoproterenol subcutaneous injection. Oral β-AR blockers treatment was performed in therapy groups. Cardiac remodeling and left ventricular function were analyzed in rats failing hearts. After 4 or 8 weeks of banding, rats developed cardiac hypertrophy and failure. Cardiac expression of ER chaperones was significantly increased. Similar to the findings above, sustained isoproterenol infusion for 2 weeks induced cardiac hypertrophy and failure with increased ER chaperones and apoptosis in hearts. β-AR blockers treatment markedly attenuated these pathological changes and reduced ER stress and apoptosis in failing hearts. On the other hand, β-AR agonist isoproterenol induced ER stress and apoptosis in cultured cardiomyocytes. β-AR blockers largely prevented ER stress and protected myocytes against apoptosis. And β-AR blockade significantly suppressed the overactivation of CaMKII in isoproterenol-stimulated cardiomyocytes and failing hearts in rats. CONCLUSIONS: Our results demonstrated that ER stress occurred in failing hearts and this could be reversed by β-AR blockade. Alleviation of ER stress may be an important mechanism underlying the therapeutic effect of β-AR blockers on heart failure.     

Stanniocalcin-1 is a naturally occurring L-channel inhibitor in cardiomyocytes: relevance to human heart failure

Cardiomyocytes of the failing heart undergo profound phenotypic and structural changes that are accompanied by variations in the genetic program and profile of calcium homeostatic proteins. The underlying mechanisms for these changes remain unclear. Because the mammalian counterpart of the fish calcium-regulating hormone stanniocalcin-1 (STC1) is expressed in the heart, we reasoned that STC1 might play a role in the adaptive-maladaptive processes that lead to the heart failure phenotype. We examined the expression and localization of STC1 in cardiac tissue of patients with advanced heart failure before and after mechanical unloading using a left ventricular assist device (LVAD), and we compared the results with those of normal heart tissue. STC1 protein is markedly upregulated in cardiomyocytes and arterial walls of failing hearts pre-LVAD and is strikingly reduced after LVAD treatment. STC1 is diffusely expressed in cardiomyocytes, although nuclear predominance is apparent. Addition of recombinant STC1 to the medium of cultured rat cardiomyocytes slows their endogenous beating rate and diminishes the rise in intracellular calcium with each contraction. Furthermore, using whole cell patch-clamp studies in cultured rat cardiomyocytes, we find that addition of STC1 to the bath causes reversible inhibition of transmembrane calcium currents through L-channels. Our data suggest differential regulation of myocardial STC1 protein expression in heart failure. In addition, STC1 may regulate calcium currents in cardiomyocytes and may contribute to the alterations in calcium homeostasis of the failing heart.     

Ca2+-ATPases in non-failing and failing heart: evidence for a novel cardiac sarco/endoplasmic reticulum Ca2+-ATPase 2 isoform (SERCA2c)

We recently documented the expression of a novel human mRNA variant encoding a yet uncharacterized SERCA [SR (sarcoplasmic reticulum)/ER (endoplasmic reticulum) Ca2+-ATPase] protein, SERCA2c [Gélébart, Martin, Enouf and Papp (2003) Biochem. Biophys. Res. Commun. 303, 676-684]. In the present study, we have analysed the expression and functional characteristics of SERCA2c relative to SERCA2a and SERCA2b isoforms upon their stable heterologous expression in HEK-293 cells (human embryonic kidney 293 cells). All SERCA2 proteins induced an increased Ca2+ content in the ER of intact transfected cells. In microsomes prepared from transfected cells, SERCA2c showed a lower apparent affinity for cytosolic Ca2+ than SERCA2a and a catalytic turnover rate similar to SERCA2b. We further demonstrated the expression of the endogenous SERCA2c protein in protein lysates isolated from heart left ventricles using a newly generated SERCA2c-specific antibody. Relative to the known uniform distribution of SERCA2a and SERCA2b in cardiomyocytes of the left ventricle tissue, SERCA2c was only detected in a confined area of cardiomyocytes, in close proximity to the sarcolemma. This finding led us to explore the expression of the presently known cardiac Ca2+-ATPase isoforms in heart failure. Comparative expression of SERCAs and PMCAs (plasma-membrane Ca2+-ATPases) was performed in four nonfailing hearts and five failing hearts displaying mixed cardiomyopathy and idiopathic dilated cardiomyopathies. Relative to normal subjects, cardiomyopathic patients express more PMCAs than SERCA2 proteins. Interestingly, SERCA2c expression was significantly increased (166+/-26%) in one patient. Taken together, these results demonstrate the expression of the novel SERCA2c isoform in the heart and may point to a still unrecognized role of PMCAs in cardiomyopathies.     

Alterations of adenylyl cyclase and G proteins in aortocaval shunt-induced heart failure

Unlike most other experimental models of congestive heart failure, the volume overload model induced by aortocaval shunt (AVS) in rats was found to exhibit enhanced beta-adrenoceptor (beta-AR) signaling. To study whether the adenylyl cyclase (AC)-G protein system is involved in such a change, we examined cardiac AC activity and protein content as well as G(s)alpha and G(i)alpha activities, protein contents, and mRNA levels in both left (LV) and right (RV) ventricles at the failing stage (16 wk after surgery). Basal and forskolin-stimulated AC activities were significantly increased in both LV and RV from the failing hearts; this change was associated with an upregulation of type V/VI AC protein. In contrast to 5'-guanylyl imidodiphosphate and NaF, the stimulatory effect of isoproterenol on AC was increased in the failing heart. Although G(s)alpha and G(i)alpha protein contents in the failing hearts were not altered, the mRNA level for G(s)alpha was decreased by 20% and that for G(i)alpha was increased by 20%. In addition, the activity of G(s)alpha, but not G(i)alpha, as assessed by toxin-catalyzed ADP ribosylation, was significantly decreased in the failing heart. Losartan and imidapril treatments improved cardiac function and attenuated alterations in mRNA levels for G(s)alpha and G(i)alpha proteins, as well as G(s)alpha activity, without affecting changes in AC protein content or activities in heart failure due to volume overload. These data suggest that increased AC activity may contribute to the enhanced beta-AR signaling in the AVS model of heart failure, whereas alterations in gene expression for G proteins may be of an adaptive nature at this stage of heart failure.     

Myocyte cytoskeletal disorganization and right heart failure in hypoxia-induced neonatal pulmonary hypertension

Previous studies have demonstrated that environmentally or genetically induced changes in the intracellular proteins that compose the cytoskeleton can contribute to heart failure. Because neonatal right ventricular myocytes are immature and are in the process of significant cytoskeletal change, we hypothesized that they may be particularly susceptible to pressure stress. Newborn calves exposed to hypobaric hypoxia (barometric pressure = 430 mmHg) for 14 days developed severe pulmonary hypertension (pulmonary arterial pressure = 101 +/- 6 vs. 27 +/- 1 mmHg) and right heart failure compared with age-matched controls. Light microscopy showed partial loss of myocardial striations in the failing neonatal right but not left ventricles and in neither ventricle of adolescent cattle dying of altitude-induced right heart failure. In neonatal calves, immunohistochemical analysis of the cytoskeletal proteins (vinculin, metavinculin, desmin, vimentin, and cadherin) showed selectively, within the failing right ventricles, patchy areas characterized by loss and disorganization of costameres and intercalated discs. Within myocytes from the failing ventricles, vinculin and desmin were observed to redistribute diffusely within the cytosol, metavinculin appeared in disorganized clumps, and vimentin immunoreactivity was markedly decreased. Western blot analysis of the failing right ventricular myocardium showed, compared with control, vinculin and desmin to be little changed in total content but redistributed from insoluble (structural) to soluble (cytosolic) fractions; metavinculin total content was markedly decreased, tubulin content increased, particularly in the structural fraction, and cadherin total content and distribution were unchanged. We conclude that hypoxic pulmonary hypertensive-induced neonatal right ventricular failure is associated with disorganization of the cytoskeletal architecture.     

Compartmentalized expression of three novel sarco/endoplasmic reticulum Ca2+ATPase 3 isoforms including the switch to ER stress, SERCA3f, in non-failing and failing human heart

The human sarco/endoplasmic reticulum (ER) Ca(2+)ATPase 3 (SERCA3) gene gives rise to SERCA3a-3f isoforms, the latter inducing ER stress in vitro. Here, we first demonstrated the co-expression of SERCA3a, -3d and -3f proteins in the heart. Evidence for endogenous proteins was obtained by using isoform-specific antibodies including a new SERCA3d-specific antibody, and either Western blotting of protein lysates or immunoprecipitation of membrane proteins. An immunolocalization study of both left ventricle tissue and isolated cardiomyocytes showed a distinct compartmentalization of the SERCA3 isoforms, as a uniform distribution of SERCA3a was detected while -3d and -3f isoforms were observed around the nucleus and in close vicinity of plasma membrane, respectively. Second, we studied their expressions in failing hearts including mixed (MCM) (n=1) and idiopathic dilated (IDCM) cardiomyopathies (n=4). Compared with controls (n=5), similar expressions of SERCA3a and -3d mRNAs were observed in all patients. In contrast, SERCA3f mRNA was found to be up-regulated in failing hearts (125+/-7%). Remarkably, overexpression of SERCA3f paralleled an increase in ER stress markers including processing of X-box-binding protein-1 (XBP-1) mRNA (176+/-24%), and expression of XBP-1 protein and glucose-regulated protein (GRP)78 (232+/-21%). These findings revisit the human heart's Ca(2+)ATPase system and indicate that SERCA3f may account for the mechanism of ER stress in vivo in heart failure.