Darier disease (DD) is an autosomal dominant disorder due to pathogenic variants of the ATP2A2 gene that causes an isolated skin manifestation based on keratinocyte disconnection and apoptosis. Systemic manifestations of DD have not been demonstrated so far, although a high incidence of neuropsychiatric syndromes suggests an involvement of the central nervous system. We report that the pathogenic ATP2A2 gene variant c.118G>A may cause cardiac involvement in patients with DD, consisting of keratinocyte and cardiomyocyte disconnection. Their common pathologic pathway, still unreported, was documented by both skin and left ventricular endomyocardial biopsies because cardiac dilatation and dysfunction appeared several decades after skin manifestations. Keratinocyte disconnection was paralleled by cardiomyocyte separation at the lateral junction. Cardiomyocyte separation was associated with cell disarray, sarcoplasmic reticulum dilatation, and increased myocyte apoptosis. Clinically, hyperkeratotic skin papules are associated with chest pain, severe muscle exhaustion, and ventricular arrhythmias that improved following administration of aminophylline, a phosphodiesterase inhibitor enhancing SERCA2 protein phosphorylation. Cardiac pathologic changes are similar to those documented in the skin, including cardiomyocyte disconnection that promotes precordial pain and cardiac arrhythmias. Phosphodiesterase inhibitors that enhance SERCA2 protein phosphorylation may substantially attenuate the symptoms.

OBJECTIVE: SERCA2, one of the P-type pumps encoded by gene ATP2A2, is the only calcium reflux channel of the endoplasmic reticulum (ER) and participates in maintaining calcium homeostasis. The present study was designed to explore SERCA2 expression pattern in auditory hair cells and the possible mechanism underlying the effects of SERCA2 on cisplatin-induced ototoxicity.
METHODS: The SERCA2 expression pattern in cochlea hair cells and HEI-OC1 cells was measured by Western blot (WB) and immunofluorescence staining. The apoptosis and its related factors were detected by TUNEL assay and WB. The expression levels of ER stress-related factors, ATF6, PERK, IRE1α, and GRP78, were measured via WB. As for the determination of SERCA2 overexpression and knockdown, plasmids and lentiviral vectors were constructed, respectively.
RESULTS: We found that SERCA2 was highly expressed in cochlea hair cells and HEI-OC1 cells. Of note, the level of SERCA2 expression in neonatal mice was remarkably higher than that in adult mice. Under the exposure of 30 μM cisplatin, SERCA2 was down-regulated significantly compared with the control group. In addition, cisplatin administration triggered the occurrence of ER stress and apoptosis. Those events were reversed by overexpressing SERCA2. On the contrary, SERCA2 knockdown could aggravate the above processes.
CONCLUSIONS: The findings from the present study disclose, for the first time, that SERCA2 is abundantly expressed in cochlea hair cells, and the suppression of SERCA2 caused by cisplatin could trigger ER homeostasis disruption, thereby implying that SERCA2 might be a promising target to prevent cisplatin-induced cytotoxicity of hair cells.

Sarcoplasmic/endoplasmic reticulum Ca2+ ATPase 2 (SERCA2) is critical in maintaining Ca2+ homeostasis. The cysteine 674 (C674) is the key redox regulatory cysteine in regulating SERCA2 activity, which is irreversibly oxidized in the renal cortex of hypertensive mice. We have reported that the substitution of C674 by serine causes SERCA2 dysfunction and increases blood pressure by induction of endoplasmic reticulum stress (ERS). This study is to explore whether the dysfunction of SERCA2 causes hypertension by interrupting mitochondrial homeostasis and inducing oxidative stress.
We used heterozygous SERCA2 C674S gene mutation knock-in (SKI) mice, where one copy of C674 was substituted by serine to represent partial C674 oxidation. In renal proximal tubule (RPT) cells, the substitution of C674 by serine decreased mitochondrial Ca2+ content, increased mitochondrial membrane potential, ATP content, and reactive oxygen species (ROS), which could be reversed by ERS inhibitor 4-phenylbutyric acid or SERCA2 agonist CDN1163. In SKI RPT cells, the redox modulator Tempol alleviated oxidative stress, downregulated the protein expression of ERS markers and soluble epoxide hydrolase, upregulated the protein expression of dopamine D1 receptor, and reduced Na+/K+- ATPase activity. In SKI mice, SERCA2 agonists CDN1163 and [6]-Gingerol, or the redox modulator Tempol increased urine output and lowered blood pressure.
The irreversible oxidation of C674 is not only an indicator of increased ROS, but also further inducing oxidative stress to cause hypertension. Activation of SERCA2 or inhibition of oxidative stress is beneficial to alleviate hypertension caused by SERCA2 dysfunction.

暂无摘要。

UNASSIGNED: Substitution of Cys674 (C674) in the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase 2 (SERCA2) causes SERCA2 dysfunction which leads to activated inositol requiring enzyme 1 alpha (IRE1α) and spliced X-box binding protein 1 (XBP1s) pathway accelerating cell proliferation of pulmonary artery smooth muscle cells (PASMCs) followed by significant pulmonary vascular remodeling resembling human pulmonary hypertension. Based on this knowledge, we intend to investigate other potential mechanisms involved in SERCA2 dysfunction-induced pulmonary vascular remodeling.
UNASSIGNED: Heterozygous SERCA2 C674S knock-in (SKI) mice of which half of cysteine in 674 was substituted by serine to mimic the partial irreversible oxidation of C674 were used. The lungs of SKI mice and their littermate wild-type mice were collected for PASMC culture, protein expression, and pulmonary vascular remodeling analysis.
UNASSIGNED: SERCA2 dysfunction increased intracellular Ca2+ levels, which activated Ca2+-dependent calcineurin (CaN) and promoted the nuclear translocation and protein expression of the nuclear factor of activated T-lymphocytes 4 (NFAT4) in an IRE1α/XBP1s pathway-independent manner. In SKI PASMCs, the scavenge of intracellular Ca2+ by BAPTA-AM or inhibition of CaN by cyclosporin A can prevent PASMC phenotypic transition. CDN1163, a SERCA2 agonist, suppressed the activation of CaN/NFAT4 and IRE1α/XBP1s pathways, reversed the protein expression of PASMC phenotypic transition markers and cell cycle-related proteins, and inhibited cell proliferation and migration when given to SKI PASMCs. Furthermore, CDN1163 ameliorated pulmonary vascular remodeling in SKI mice.
UNASSIGNED: SERCA2 dysfunction promotes PASMC phenotypic transition and pulmonary vascular remodeling by multiple mechanisms, which could be improved by SERCA2 agonist CDN1163.
‘What is already known’ l The dysfunction of SERCA2 promotes PASMC hyperproliferation and pulmonary vascular remodeling through activation of the IRE1α/XBP1s pathway.‘What this study adds’ l The dysfunction of SERCA2 activates the Ca2+-dependent CaN-mediated NFAT4 pathway to promote the PASMC phenotypic transition.l Revitalization of SERCA2 suppresses PASMC phenotypic transition and pulmonary vascular remodeling caused by SERCA2 dysfunction.‘Clinical significance’ l SERCA2 dysfunction-induced pulmonary vascular remodeling involves more than one mechanism, implicating that more drugable targets are to be discovered.l SERCA2 is a potential therapeutic target for preventing pulmonary vascular remodeling.

The endoplasmic reticulum (ER) stores large amounts of calcium (Ca2+), and the controlled release of ER Ca2+ regulates a myriad of cellular functions. Although altered ER Ca2+ homeostasis is known to induce ER stress, the mechanisms by which ER Ca2+ imbalance activate ER stress pathways are poorly understood. Stromal-interacting molecules STIM1 and STIM2 are two structurally homologous ER-resident Ca2+ sensors that synergistically regulate Ca2+ influx into the cytosol through Orai Ca2+ channels for subsequent signaling to transcription and ER Ca2+ refilling. Here, we demonstrate that reduced STIM2, but not STIM1, in colorectal cancer (CRC) is associated with poor patient prognosis. Loss of STIM2 causes SERCA2-dependent increase in ER Ca2+, increased protein translation and transcriptional and metabolic rewiring supporting increased tumor size, invasion, and metastasis. Mechanistically, STIM2 loss activates cMyc and the PERK/ATF4 branch of ER stress in an Orai-independent manner. Therefore, STIM2 and PERK/ATF4 could be exploited for prognosis or in targeted therapies to inhibit CRC tumor growth and metastasis.

Cell-penetrating peptides (CPPs) are short peptide sequences that have the ability to cross the cell membrane and deliver cargo. Although it is critical that CPPs accomplish this task with minimal off-target effects, such actions have in many cases not been robustly screened. We presently investigated whether the commonly used CPPs TAT and the polyarginines Arg9 and Arg11 exert off-target effects on cellular Ca2+ homeostasis. In experiments employing myocytes and homogenates from the cardiac left ventricle or soleus muscle, we observed marked inhibition of Ca2+ recycling into the sarcoplasmic reticulum (SR) following incubation with polyarginine CPPs. In both tissues, the rate of SR Ca2+ leak remained unchanged, indicating that protracted Ca2+ removal from the cytosol stemmed from inhibition of the SR Ca2+ ATPase 2 (SERCA2). No such inhibition occurred following treatment with TAT, or in preparations from the SERCA1-expressing extensor digitorum longus muscle. Experiments in HEK cells overexpressing individual SERCA isoforms confirmed that polyarginine incubation specifically inhibited the activity of SERCA2a and 2b, but not SERCA1 or 3. The attenuation of SERCA2 activity was not dependent on the presence of phospholamban, and ELISA-based analyses rather revealed direct interaction between the polyarginines and the actuator domain of the protein. Surface plasmon resonance experiments confirmed strong binding within this region of SERCA2, and slow dissociation between the two species. Based on these observations, we urge caution when employing polyarginine CPPs. Indeed, as SERCA2 is expressed in diverse cell types, the wide-ranging consequences of SERCA2 binding and inhibition should be anticipated in both experimental and therapeutic settings.

The well-known insulin-like growth factor 1 (IGF1)/IGF-1 receptor (IGF-1R) signaling pathway is overexpressed in many tumors, and is thus an attractive target for cancer treatment. However, results have often been disappointing due to crosstalk with other signals. Here, we report that IGF-1R signaling stimulates the growth of hepatocellular carcinoma (HCC) cells through the translocation of IGF-1R into the ER to enhance sarco-endoplasmic reticulum calcium ATPase 2 (SERCA2) activity. In response to ligand binding, IGF-1Rβ is translocated into the ER by β-arrestin2 (β-arr2). Mass spectrometry analysis identified SERCA2 as a target of ER IGF-1Rβ. SERCA2 activity is heavily dependent on the increase in ER IGF-1Rβ levels. ER IGF-1Rβ phosphorylates SERCA2 on Tyr990 to enhance its activity. Mutation of SERCA2-Tyr990 disrupted the interaction of ER IGF-1Rβ with SERCA2, and therefore ER IGF-1Rβ failed to promote SERCA2 activity. The enhancement of SERCA2 activity triggered Ca2+ER perturbation, leading to an increase in autophagy. Thapsigargin blocked the interaction between SERCA2 and ER IGF-1Rβ and therefore SERCA2 activity, resulting in inhibition of HCC growth. In conclusion, the translocation of IGF-1R into the ER triggers Ca2+ER perturbation by enhancing SERCA2 activity through phosphorylating Tyr990 in HCC.

Termination codon readthrough (TCR) is a process in which ribosomes continue to translate an mRNA beyond a stop codon generating a C-terminally extended protein isoform. Here, we demonstrate TCR in mammalian NNAT mRNA, which encodes NNAT, a proteolipid important for neuronal differentiation. This is a programmed event driven by cis-acting RNA sequences present immediately upstream and downstream of the canonical stop codon and is negatively regulated by NONO, an RNA-binding protein known to promote neuronal differentiation. Unlike the canonical isoform NNAT, we determined that the TCR product (NNATx) does not show detectable interaction with the sarco/endoplasmic reticulum Ca2+-ATPase isoform 2 Ca2+ pump, cannot increase cytoplasmic Ca2+ levels, and therefore does not enhance neuronal differentiation in Neuro-2a cells. Additionally, an antisense oligonucleotide that targets a region downstream of the canonical stop codon reduced TCR of NNAT and enhanced the differentiation of Neuro-2a cells to cholinergic neurons. Furthermore, NNATx-deficient Neuro-2a cells, generated using CRISPR-Cas9, showed increased cytoplasmic Ca2+ levels and enhanced neuronal differentiation. Overall, these results demonstrate regulation of neuronal differentiation by TCR of NNAT. Importantly, this process can be modulated using a synthetic antisense oligonucleotide.

Gonnot et al. [1] thoroughly investigated the regulatory role of glycogen synthase kinase 3 beta (GSK3β) in modulating cardiac isoform 2 of sarcoplasmic/endoplasmic reticulum Ca2+ ATPase (SERCA2) activity. They have found that in ischemic hearts of patients and mouse-GSK3β -mediated SERCA2 phosphorylation at serine 663 dampens the SERCA2 pump activity and induces Ca2+ overload which sensitizes towards myocardial ischemia-reperfusion (I/R) injury. The inhibition of serine 663 phosphorylation significantly increases SERCA2 activity and, by preventing cytosolic and mitochondrial Ca2+ overload, reduces cell death during reperfusion. Augmented SERCA2 activity also substantially improves excitation-contraction coupling in cardiomyocytes upon recovery from reperfusion injury. This study provides valuable insights into pathophysiological relevance of GSK3β -mediated SERCA2 phosphorylation in the context of heart diseases and paves the way for designing novel clinical therapeutic approaches to alleviate post infartion heart failure.