Orai1 plays a critical role in Orai3 synthesis and stability in luminal breast cancer cells.

Tumor microenvironment substantially influences the process of tumorigenesis. Extracellular acidification within the tumor microenvironment is an indicator of an aggressive cancer and a marker for poor patient outcome. In solid tumors, hypoxia leads to extracellular acidosis. Carbonic anhydrases (CA) are thought to regulate intracellular and extracellular pH (pHi and pHe, respectively). To explore the effect of CAs in breast cancer, we compared the expression and activity of two membrane bound CAs, CAIX and CAXII, between triple negative breast cancer cells (TNBCs) and luminal breast cancer cells (LBCs). We chose five different TNBC and LBC lines. Our data show that, among the TNBC lines, CAIX expression increased in three of the five lines: HBL100, SUM159, and the new UFH-001 cells under hypoxic condition. UFH-001 cells also showed strong constitutive expression. None of these TNBC lines expressed CAXII or estrogen receptor (ER). In LBC lines, four of the five lines constitutively expressed CAXII: T47D, MCF7, SKBR and SUM52 cells. CAXII expression was not hypoxia-dependent. Each of the five luminal lines expressed ER. We also examined CA expression in a tumor graft model. In tumors grown from cells derived from TNBC patients, we observed CAIX expression in four of six sample sets. In tumors derived from ER-positive LBC patients, all five expressed CAXII. We also used the 18O exchange method to assess CA activity. Two TNBC lines: UFH-001 and HBL100 cells showed that CAIX activity increased in hypoxic conditions which was blocked by an impermeant sulfonamide CA inhibitor (N3500). In the luminal lines, we detected CAXII activity in T47D and MCF7 cells that was also inhibited by N3500. Like CAXII protein expression in these cells, CAXII activity was not affected by hypoxia. We also evaluated the effect of pH on CA activity in TNBC and LBC lines. Both CAIX and CAXII showed increased activity in response to reduced pH, which is expected in a bicarbonate-based system. However, UFH-001 cells also exhibited a hypoxic-dependent increase in CAIX activity which is associated with increased protein expression. In conclusion, these observations demonstrate that CAIX expression is associated with the TNBC phenotype. Based on our activity data, we would predict that CA activity in TNBC tumors will be sensitive to both hypoxia (based on enhanced expression) and reduced pH. This change in activity may serve to regulate pH in the tumor microenvironment favoring an aggressive phenotype. On the other hand, LBC tumors, which are ER-positive, are only associated with CAXII expression. In luminal cells, we expect that only pH and not hypoxia will affect CAXII activity. This may, in part, explain the more positive prognosis in patients with CAXII expression. Note: This abstract was not presented at the meeting. Citation Format: Zhijuan Chen, Mam Y. Mboge, Chingkuang Tu, Lingbao Ai, Coy Heldermon, Susan C. Frost. Comparison of carbonic anhydrase & activity between triple-negative & luminal breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5936. doi:10.1158/1538-7445.AM2017-5936

Store-operated Ca(2+) entry (SOCE) encoded by Orai1 proteins is a ubiquitous Ca(2+)-selective conductance involved in cellular proliferation and migration. We recently described up-regulation of Orai3 channels that selectively mediate SOCE in estrogen receptor α-expressing (ERα(+)) breast cancer cells. However, the connection between ERα and Orai3 and the role of Orai3 in tumorigenesis remain unknown. Here, we show that ERα knockdown decreases Orai3 mRNA (by ∼63%) and protein (by ∼44%) with no effect on Orai1. ERα knockdown decreases Orai3-mediated SOCE (by ∼43%) and the corresponding Ca(2+) release-activated Ca(2+) (CRAC) current (by ∼42%) in ERα(+) MCF7 cells. The abrogation of SOCE in MCF7 cells on ERα knockdown can be rescued by ectopic expression of Orai3. ERα activation increased Orai3 expression and SOCE in MCF7 cells. Epidermal growth factor (EGF) and thrombin stimulate Ca(2+) influx into MCF7 cells through Orai3. Orai3 knockdown inhibited SOCE-dependent phosphorylation of extracellular signal-regulated kinase (ERK1/2; by ∼44%) and focal adhesion kinase (FAK; by ∼46%) as well as transcriptional activity of nuclear factor for activated T cells (NFAT; by ∼49%). Significantly, Orai3 knockdown selectively decreased anchorage-independent growth (by ∼58%) and Matrigel invasion (by ∼44%) of ERα(+) MCF7 cells with no effect on ERα(-) MDA-MB231 cells. Moreover, Orai3 knockdown inhibited ERα(+) cell tumorigenesis in immunodeficient mice (∼66% reduction in tumor volume). These data establish Orai3 as an ERα-regulated channel and a potential selective therapeutic target for ERα(+) breast cancers.

Intracellular Ca2+ signalling is essential to the regulation of many biological mechanisms, which have distinct functional roles. The promiscuity of Ca2+ signalling represents a challenge for fine signalling transduction, and this appears particularly evident in somatosensory neurons, which are capable of generating and relaying signal related to well-defined, but discrete, sensory information, yet express numerous Ca2+ channels. To overcome this specificity issue, Ca2+ signalling is often restricted to subcellular compartments (nanodomains) – specialised regions within a cell where Ca2+ undergoes local modulation, allowing fine spatiotemporal tuning. Endoplasmic reticulum (ER)-plasma membrane (PM) junctions are an example of such nanodomains, where physical proximity between these two membranes is critical for store operated Ca2+ entry (SOCE). In mammalian cells, SOCE complexes have of two main components: (1) the ER Ca2+ sensor stromal interaction protein 1 (STIM1), and (2) the pore-forming subunits of the Ca2+ release-activated Ca2+ channel (CRAC) protein 1–3 (Orai1-3). Activation of heterotrimeric Gq coupled receptor triggers phospholipase C enzyme activation, subsequent formation of inositol,1,4,5, triphosphate (IP3), and release of Ca2+ from the ER; this leads to a drop in ER Ca2+ levels, which leads to STIM1 oligomerisation. Finally, via protein-protein interaction, STIM1 opens Orai1 channels on the plasma membrane, allowing SOCE (Woo et al. 2018). Various adaptor proteins are required to stabilize PM-ER junctions and to allow SOCE, among which are junctophilins (JPH). The JPHs are a family of junctional membrane complex-associated proteins whose C- and N-termini are situated in the PM and ER, respectively. Thus, they form junctional complexes between these two membrane compartments. There are four junctophilin isoforms (JPH1–4) displaying different tissue specificity of expression. While JPH3–4 are ubiquitously expressed in the central nervous system (CNS), JPH expression in dorsal root ganglia (DRG) has not yet been explored in depth. ER-PM junctions exist in DRG somatosensory neurons, and functional SOCE complexes have also been reported in these neurons (Gemes et al. 2011). SOCE has been associated with pain elicited by pro-inflammatory mediators such as bradykinin (BK). BK acts via its B2 receptors (B2R) to excite peripheral neurons by inhibiting M-type K+ channels and activating Ca2+-dependent Cl– channels (Brown and Passmore, 2010) (Liu et al. 2010). The role of junctophilins in this process has not previously been elucidated. In a recent publication in The Journal of Physiology, Hogea et al. (2021) characterised the role of junctophilins in SOCE in DRG somatosensory neurons and investigated their potential involvement in inflammatory pain in vitro and in vivo. The authors first aimed to identify which isoforms of the JPH, STIM and Orai families were expressed in primary sensory neurons. Utilising immunohistochemistry and western blot analyses, they found that three JPH isoforms – JPH1, JPH3 and JPH4 – were expressed in neonatal male rat DRG neurons. JPH4 was the predominant isoform, found to be expressed in 76% of analysed DRG neurons. It was significantly more abundant than the other isoforms and seemingly preferentially located at the PM, prompting the group to focus on this isoform for further experiments. Immunohistochemistry also confirmed the expression of STIM1-2, as well as Orai1-3 in rat DRG neurons, with STIM 1 and Orai1 being the predominant isoforms, expressed in 54% and 72% of DRG neurons, respectively. Interestingly, the authors showed that upon ER Ca2+ store depletion, JPH4 highly co-localised with STIM1 and Orai1, essential for SOCE in DRG neurons. JPH4, STIM1 and Orai1 were found to be expressed uniformly across neurons of different somatic sizes. JPH4 expression was found in 75% and 84% of neurons labelled for NF200 and TRPV1, respectively. It should be noted that not all polymodal nociceptors express TRPV1, and furthermore TRPV1 is also expressed in some Aδ fibres. This overlap was not addressed and other markers for different sensory neuron modalities were not used in this study. Moving forward, the group investigated the role of JPH4 in GPCR activation-mediated store depletion in primary somatosensory neurons. Using proximity ligation assay and super-resolution imaging, they confirmed co-localization of JPH4 with STIM1 and Orai1, and that – similarly to the established clustering of STIM1 and Orai1 – ER Ca2+ store depletion stimulated recruitment of JPH4 to this cluster as well. Immunohistochemical and western blot analyses following siRNA-mediated JPH4 knockdown revealed that JPH4 was in fact necessary for efficient clustering of STIM1 and Orai1 after BK treatment. This is in agreement with previous literature showing JPH1–2 dependence of STIM1-Orai1 ER clustering in skeletal myocytes and CNS (Takeshima et al. 2015). To clarify JPH4 role in Ca2+ signalling, the authors performed a set of in vitro calcium imaging experiments. First, they demonstrated that SOCE could be measured in cultured DRG neurons. Stimulating DRGs with BK in the absence of extracellular Ca2+ induced a robust increase in cytoplasmic Ca2+, depleting ER stores; Ca2+ was then added back to the extracellular solution, inducing a second Ca2+ transient – this was prevented by treating DRGs with SOCE inhibitors, indicating that the increase in cytoplasmic Ca2+ during the Ca2+ add-back involved the SOCE complex. Next, they showed that in JPH4-silenced neurons the initial BK-induced Ca2+ transient was maintained, but the increase in cytoplasmic Ca2+ was significantly attenuated when Ca2+ was added back to the extracellular solution, suggesting that JPH4 plays a functional role in SOCE. The authors then questioned if this apparent JPH4 involvement in replenishing ER Ca2+ stores would affect recurrent GPCR-mediated Ca2+ signalling. Since prolonged or repeated GPCR stimulation can lead to decreased responsiveness to an agonist treatment, the ER Ca2+ stores were first depleted via different GPCRs – purinergic P2Y receptors – by stimulating DRG neurons with the P2Y agonist ATP in the absence of extracellular Ca2+. Subsequently, Ca2+ was added back to the extracellular solution, and the neurons were stimulated with BK; interestingly, the release of Ca2+ from the ER in response to BK was attenuated by 40% in JPH4-silenced neurons. These experiments suggest that not only JPH4 is required for STIM1-Orai1 clustering upon ER Ca2+ store depletion, but that by impairing SOCE, JPH4 knockdown ultimately decreases DRG sensitivity during repeated GPCR stimulation. Finally, to determine whether JPH4 is involved in pain perception, Hogea et al. (2021) delivered cholesterol-conjugated JPH4 siRNA to the lumbar region (L5-L6) of adult rats to silence JPH4 gene expression and performed behavioural experiments. Intraplantar injection of BK induces inflammation and nocifensive behaviour in rodents. The authors demonstrated that while normal mechanical and thermal sensitivity was maintained in rats with decreased JPH4 expression, the time-span of nocifensive behaviour significantly decreased in these animals. Overall, this experiment corroborates the molecular findings, suggesting that JPH4 is implicated in peripheral BK-mediated inflammatory pain. Whilst the authors present strong evidence of their hypothesis regarding the involvement of JPH4 in inflammatory mechanisms at PM-ER nanodomains, the use of other in vivo models such as knockout mice or the inclusion of female animals could have strengthened these findings and clarified whether these mechanisms are conserved across the two sexes or in different species. Moreover, whilst DRG cells bodies are of growing interest in the context of nociception input regulation, it would have been valuable to investigate these JPH4-mediated mechanisms at nerve terminals – the loci of inflammatory excitation. However, given (1) the presence of functional BK receptors in the periphery as demonstrated by BK-induced pain response in vivo, (2) the expression of ER in proximal C-fibres, and (3) the reduction of duration of nocifensive behaviours in vivo upon JPH4 knockdown in rats’ lumbar region, it is reasonable to speculate that the observed effects are also relevant for the peripheral portions of somatosensory nerves. To conclude, the work of Hogea et al. (2021) has successfully provided the first evidence of the role of the junctophilin isoform JPH4 in SOCE and inflammatory pain mediation in peripheral somatosensory neurons of male rats. No competing interests declared. A.F. and G.T.: conception or design of the work; drafting the work or revising it critically for important intellectual content; final approval of the version to be published; agreement to be accountable for all aspects of the work. All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed. None. We thank Dr Viktor Lukacs for the assistance in writing this article.

Piper chaba (Piperaceae) is a medicinal spice plant that possesses several pharmacological activities. In the present study, we for the first time studied the effect of P. chaba extract on breast cancer cells. P. chaba stem methanolic (PCSM) extract produced time and dose dependent cytotoxicity in luminal breast cancer cells (MCF-7 and T47D) with a minimal toxicity in breast normal cells (MCF-10A) at 10-100 µg/mL concentration. PCSM extract exerts 16.79 and 31.21 µg/mL IC50 for T47D and MCF-7 cells, respectively, in 48 h treatment. PCSM significantly arrests the T47D cells at the G0/G1 phase by reducing the CCND1 and CDK4 expression at mRNA and protein levels. PCSM extract treatment significantly altered nuclear morphology, mitochondria membrane potential, and production of reactive oxygen species in T47D cells at IC50 concentration. Extract treatment significantly altered the Bax/Bcl-2 ratio and altered caspase 8 and 3 mRNA/protein levels in T47D cells. Confocal microscopy showed an increase in late apoptosis in PCSM extract-treated breast cancer cells at IC50 . Further, an increased caspase 9 and caspase 3/7 enzymatic activity was observed in test cells compared with nontreated cells. In conclusion, P. chaba phytocompound possesses the potential to induce cell cycle arrest and induce apoptosis in luminal breast cancer cells.

Store-operated Ca²⁺ entry (SOCE) is a major pathway for Ca²⁺ entry that regulates several cellular functions. SOCE remodeling mediated by changes in the expression and/or function of the Orai channels results in the reorganization of intracellular Ca2+ homeostasis leading to a variety of pathologies, including cancer. Notably, a significant alteration of Orai function has been reported in breast cancer cells, where the dysregulation of the Notch1 signaling pathway plays a role in the development and progression of cancer hallmarks. Here, we have investigated the possible role of Notch1 in the regulation of the expression of Orai1 and Orai3 in different breast cancer cell lines. Expression of the active form of Notch1, as well as cell stimulation with the Notch1 agonist Jagged-1 (Jag-1), demonstrates a differential role of Notch1 in the regulation of Orai expression in non-tumoral breast epithelial cells and triple negative or luminal breast cancer cells. The role of Notch1 was confirmed using DAPT, a γ-secretase inhibitor that prevents activation of the Notch pathway. Modulation of Orai1 and Orai3 expression by Notch1 was paralleled by changes in SOCE. The effect in Orai expression mediated by activation of Notch1 signaling pathway was mimicked by the expression of HEY1 or the non-phosphorylatable HEY1-S68A mutant; by contrast, expression of the phosphomimetic HEY1-S68D mutant was without effect on Orai expression. Understanding the Notch1-HEY1-Orai axis might provide insights into the development of subtype-specific therapeutic strategies targeting breast cancer.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-33071-x.

Undertaker, a Drosophila Junctophilin, Links Draper-Mediated Phagocytosis and Calcium Homeostasis

Store-operated Ca2+ entry (SOCE) is an important Ca2+ influx pathway, which is defined by the fact that depletion of intracellular Ca2+ stores, mainly the endoplasmic reticulum (ER), triggers the opening of Ca2+ channels in the plasma membrane. The best characterized SOC channel is the Ca2+ release-activated Ca2+ (CRAC) channel, which was first described in cells of the immune system but has since been reported in many different cell types. CRAC channels are multimers of ORAI family proteins, of which ORAI1 is the best characterized. They are activated by stromal interaction molecules (STIM) 1 and 2, which respond to the depletion of intracellular Ca2+ stores with oligomerization and binding to ORAI proteins. The resulting SOCE is critical for the physiological function of many cell types including immune cells and platelets. Recent studies using cell lines, animal models and primary cells from human patients with defects in SOCE have highlighted the importance of this Ca2+ entry mechanism in a variety of pathophysiological processes. This review focuses on the role of SOCE in immunity to infection, allergy, hemostasis and cancer.

In malignant hyperthermia (MH), mutations in RyR1 underlie direct activation of the channel by volatile anesthetics, leading to muscle contracture and a life-threatening increase in core body temperature. The aim of the present study was to establish whether the associated depletion of sarcoplasmic reticulum (SR) Ca(2+) triggers sarcolemmal Ca(2+) influx via store-operated Ca(2+) entry (SOCE). Samples of vastus medialis muscle were obtained from patients undergoing assessment for MH susceptibility using the in vitro contracture test. Single fibers were mechanically skinned, and confocal microscopy was used to detect changes in [Ca(2+)] either within the resealed t-system ([Ca(2+)](t-sys)) or within the cytosol. In normal fibers, halothane (0.5 mM) failed to initiate SR Ca(2+) release or Ca(2+)(t-sys) depletion. However, in MH-susceptible (MHS) fibers, halothane induced both SR Ca(2+) release and Ca(2+)(t-sys) depletion, consistent with SOCE. In some MHS fibers, halothane-induced SR Ca(2+) release took the form of a propagated wave, which was temporally coupled to a wave of Ca(2+)(t-sys) depletion. SOCE was potently inhibited by "extracellular" application of a STIM1 antibody trapped within the t-system but not when the antibody was denatured by heating. In conclusion, (i) in human MHS muscle, SR Ca(2+) depletion induced by a level of volatile anesthetic within the clinical range is sufficient to induce SOCE, which is tightly coupled to SR Ca(2+) release; (ii) sarcolemmal STIM1 has an important role in regulating SOCE; and (iii) sustained SOCE from an effectively infinite extracellular Ca(2+) pool may contribute to the maintained rise in cytosolic [Ca(2+)] that underlies MH.

Extended synaptotagmins (E-Syts) are endoplasmic reticulum (ER)-associated proteins that facilitate the tethering of the ER to the plasma membrane (PM), participating in lipid transfer between the membranes and supporting the Orai1-STIM1 interaction at ER-PM junctions. Orai1 and STIM1 are the core proteins of store-operated Ca2+ entry (SOCE), a major mechanism for Ca2+ influx that regulates a variety of cellular functions. Aberrant modulation of SOCE in cells from different types of cancer has been reported to underlie the development of several tumoral features. Here we show that estrogen receptor-positive (ER+) breast cancer MCF7 and T47D cells and triple-negative breast cancer (TNBC) MDA-MB-231 cells overexpress E-Syt1 and E-Syt2 at the protein level; the latter is also overexpressed in the TNBC BT20 cell line. E-Syt1 and E-Syt2 knockdown was without effect on SOCE in non-tumoral MCF10A breast epithelial cells and ER+ T47D breast cancer cells; however, SOCE was significantly attenuated in ER+ MCF7 cells and TNBC MDA-MB-231 and BT20 cells upon transfection with siRNA E-Syt1 or E-Syt2. Consistent with this, E-Syt1 and E-Syt2 knockdown significantly reduced cell migration and viability in ER+ MCF7 cells and the TNBC cells investigated. To summarize, E-Syt1 and E-Syt2 play a relevant functional role in breast cancer cells.

Background: Melanoma has poor prognosis due to its strong metastatic ability that is mainly controlled by cell migration. Store operated Ca2+ entry (SOCE), which is defined as Ca2+ entry from extracellular space triggered with depletion of Ca2+ store in the endoplasmic reticulum (ER), largely regulates Ca2+ homeostasis in non-excitable cells. However, little is known about the role of SOCE in melanoma. Here we report that expressions of SOCE modulators, i.e., ORAI calcium release-activated calcium modulator (Orai), which is the plasma membrane Ca2+ channel, and stromal interaction molecule (STIM), an activator of Orai in the ER, in cultured melanoma cells and human melanoma tissues. We also examined the effect of these modulators on SOCE and cell migration in melanoma cells Method: Western blot analysis in SK-Mel-2 cells, and immunohistochemistry in human tissue microarray, were performed to examine expressions of Orai1 or STIM1. Intracellular Ca2+ was measured with fluo-4 AM, Ca2+-sensing fluorescent dye, in human metastatic melanoma cell lines (SK-MEL-2, SK-MEL-24 and C8161 cells), a mouse melanoma cell line (B16 cells), a human melanocyte cell line, (HEMA-LP), and a mouse melanocyte cell line (Melan-A). Cell migration was examined with the Boyden chambers. In order to ablate Orai1 or STIM1, shRNA for each protein was induced in SK-Mel-2 cells with lentiviral infection. Results: Expressions of Orai1 and STIM1 were observed in melanoma cells by western blot, and in human melanoma tissues by immunohistochemistry. SOCE, as demonstrated by enhancement of Ca2+ enhancement after Ca2+ depletion with thapsigargin, was clearly observed in melanoma cell lines, but not in melanocytes, suggesting that SOCE is enhanced in melanoma cells compared to melanocytes. When Orai1 was ablated in SK-MEL-2 cells, the peak Ca2+ signal within SOCE was reduced by 75 % compared to control (2.78±0.44 (control shRNA) vs. 1.44±0.18 (Orai1 shRNA) F/F0 ratio normalized to baseline, p<0.05, n=5). Similarly, ablation of STIM1 reduces the peak Ca2+ signal within SOCE by 37 % compared to control (2.78±0.44 (control shRNA) vs. 2.12±0.63 (STIM1 shRNA) F/F0 ratio normalized to baseline, F/F0, normalized to baseline, p<0.05, n=10). These data suggest that Orai1 and STIM1 are involved in SOCE in melanoma cells. We next examined the role of Orai1 and STIM1 in melanoma cell migration. Ablation of Orai1 reduced cell migration by 46 % (48±4.6 (control shRNA) vs. 26±3.2 (Orai1 shRNA) cells/field, p<0.05, n=4). Similarly, ablation of STIM1 decreased cell migration by 40 % (48±4.6 (control shRNA) vs. 29±4.8 (STIM1 shRNA) cells/field, p<0.05, n=4). These data suggest that Orai1 and/or STIM1 are involved in basal cell migration in melanoma. Conclusion: SOCE, which is regulated by Orai1 and STIM1 in melanoma cells, plays a major role in Ca2+ homeostasis, and cell migration in melanoma, and thus potentially metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5259. doi:10.1158/1538-7445.AM2011-5259

NOX4 alleviates breast cancer cell aggressiveness by co-ordinating mitochondrial turnover through PGC1α/Drp1 axis

To investigate the differences in the expression profiles of circular RNA (circRNA) between luminal breast cancer cells and normal breast cells. Total RNA extracted from luminal breast cancer cells MCF7 and normal breast cells MCF10A was digested with Rnase R to remove linear RNAs and enrich circRNAs. The enriched circRNAs were amplified and transcribed into fluorescent cRNAs using a random priming method, and were hybridized onto the circRNA hybridization array. The circRNA expression profiles of MCF7 and MCF10A cells were analyzed using Agilent Feature Extraction software. Quantile normalization and subsequent data processing were performed, and volcano plot filtering and hierarchical clustering were utilized to analyze the circRNA expression patterns. The expressions of 3 circRNAs with significant log fold changes were validated using qPCR. The hybridization array data revealed significant differences in the circRNA expression profiles between MCF7 and MCF10A cells. Compared with those of MCF10A cells, the 12910 circRNAs expressed in MCF7 cells showed 5964 up-regulated, 81 consistently regulated, and 6865 down-regulated circRNAs; 343 circRNAs showed a log fold change by more than 2 folds, among which 213 circRNAs were up-regulated and 130 were down-regulated. Nine circRNAs showed differential expressions by more than 2 folds, including 8 up-regulated ones, namely hsa_circRNA_061260 (6.02 folds), hsa_circRNA_103933 (5.96 folds), hsa_circRNA_005239 (5.84 folds), hsa_circRNA_100689 (5.69 folds), hsa_circRNA_004087 (5.60 folds), hsa_circRNA_104420 (5.25 folds), hsa_circRNA_104421 (5.13 folds) and hsa_circRNA_101222 (5.03 folds); only one circRNA was down-regulated, namely hsa_circRNA_104864 (5.09 folds). The expressions of hsa_circRNA_100689, hsa_circRNA_005239 and hsa_circRNA_104864 were further validated by qPCR, which yielded consistent results with the microarray data. The circRNA expression profiles differ significantly between luminal breast cancer cells and normal breast cells. These differentially expressed circRNAs may serve as potential novel targets for the diagnosis of luminal breast cancer.

Stim1 and Orai1 are ubiquitous proteins that have long been known to mediate Ca2+ release-activated Ca2+ (CRAC) current (ICRAC) and store-operated Ca2+ entry (SOCE) only in non-excitable cells. SOCE is activated following the depletion of the endogenous Ca2+ stores, which are mainly located within the endoplasmic reticulum (ER), to replete the intracellular Ca2+ reservoir and engage specific Ca2+-dependent processes, such as proliferation, migration, cytoskeletal remodeling, and gene expression. Their paralogs, Stim2, Orai2 and Orai3, support SOCE in heterologous expression systems, but their physiological role is still obscure. Ca2+ inflow in neurons has long been exclusively ascribed to voltage-operated and receptor-operated channels. Nevertheless, recent work has unveiled that Stim1–2 and Orai1-2, but not Orai3, proteins are also expressed and mediate SOCE in neurons. Herein, we survey current knowledge about the neuronal distribution of Stim and Orai proteins in rodent and human brains; we further discuss that Orai2 is the main pore-forming subunit of CRAC channels in central neurons, in which it may be activated by either Stim1 or Stim2 depending on species, brain region and physiological stimuli. We examine the functions regulated by SOCE in neurons, where this pathway is activated under resting conditions to refill the ER, control spinogenesis and regulate gene transcription. Besides, we highlighted the possibility that SOCE also controls neuronal excitation and regulate synaptic plasticity. Finally, we evaluate the involvement of Stim and Orai proteins in severe neurodegenerative and neurological disorders, such as Alzheimer’s disease and epilepsy.

ORAI1 mutations abolishing store-operated Ca2+ entry cause anhidrotic ectodermal dysplasia with immunodeficiency

FOXF2 oppositely regulates stemness in luminal and basal-like breast cancer cells through the Wnt/beta-catenin pathway