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Tumor metastasis is the primary cause of treatment failure and cancer-related deaths. Store-operated Ca2+ entry (SOCE), which is mediated by stromal interaction molecules (STIM) and ORAI proteins, has been implicated in the tumor invasion-metastasis cascade. Epithelial-mesenchymal transition (EMT) is a cellular program that enables tumor cells to acquire the capacities needed for migration and invasion and the formation of distal metastases. Tumor-associated angiogenesis contributes to metastasis because aberrantly developed vessels offer a path for tumor cell dissemination as well as supply sufficient nutrients for the metastatic colony to develop into metastasis. Recently, increasing evidence has indicated that SOCE alterations actively participate in the multi-step process of tumor metastasis. In addition, the dysregulated expression of STIM/ORAI has been reported to be a predictor of poor prognosis. Herein, we review the latest advances about the critical role of SOCE in the tumor metastasis cascade and the underlying regulatory mechanisms. We emphasize the contributions of SOCE to the EMT program, tumor cell migration and invasion, and angiogenesis. We further discuss the possibility of modulating SOCE or intervening in the downstream signaling pathways as a feasible targeting therapy for cancer treatment.

Animal behavior was classically considered to be determined exclusively by neuronal activity, whereas surrounding glial cells such as astrocytes played only supportive roles. However, astrocytes are as numerous as neurons in the mammalian brain, and current findings indicate a chemically based dialog between astrocytes and neurons. Activation of astrocytes by synaptically released neurotransmitters converges on regulating intracellular Ca2+ in astrocytes, which then can regulate the efficacy of near and distant tripartite synapses at diverse timescales through gliotransmitter release. Here, we discuss recent evidence on how diverse behaviors are impacted by this dialog. These recent findings support a paradigm shift in neuroscience, in which animal behavior does not result exclusively from neuronal activity but from the coordinated activity of both astrocytes and neurons. Decoding how astrocytes and neurons interact with each other in various brain circuits will be fundamental to fully understanding how behaviors originate and become dysregulated in disease.

Studies from rabbit show that anoctamin-1 (Ano1) channels expressed in urethral interstitial cells of Cajal (ICCs) serve as a source of depolarization for smooth muscle cells, increasing excitability and tone. However, the role of urethral Ano1 channels is unclear, owing to differences in the species examined and experimental approaches. We summarize findings from different species on the role of urethral ICC and Ano1 channels in urethral contractility and outline proposals for clarifying this topic using cell-specific optogenetic approaches.

Although cultured adult cardiac myocytes in combination with cell-level genetic modifications have been adopted for the study of protein function, the cellular alterations caused by the culture conditions themselves need to be clarified before we can interpret the effects of genetically altered proteins. We systematically compared the cellular morphology, global Ca2+ signaling, elementary Ca2+ release (sparks), and arrangement of ryanodine receptor (RyR) clusters in short-term (2 days)-cultured adult rat ventricular myocytes with those of freshly isolated myocytes. The transverse (t)-tubules were remarkably decreased (to ∼25%) by culture, and whole-cell capacitance was reduced by ∼35%. The magnitude of depolarization-induced Ca2+ transients decreased to ∼50%, and Ca2+ transient decay was slowed by culture. The culture did not affect sarcoplasmic reticulum (SR) Ca2+ loading. Therefore, fractional Ca2+ release was attenuated by culture. In the cultured cells, the L-type Ca2+ current (ICa) was smaller (∼50% of controls) and its inactivation was slower. In cultured myocytes, there were significantly fewer (∼50% of control) Ca2+ sparks, the local Ca2+ releases through RyR clusters, compared with in freshly isolated cells. Amplitude and kinetics (duration and time-to-peak) of individual sparks were similar, but they showed greater width in cultured cells. Immunolocalization analysis revealed that the cross-striation of RyRs distribution became weaker and less organized, and that the density of RyR clusters decreased in cultured myocytes. Our data suggest that the loss of t-tubules and generation of compromised Ca2+ transients and ICa in short-term adult ventricular cell culture are independent of SR Ca2+ loading status. In addition, the deteriorated arrangement of the RyR-clusters and their decreased density after short-term culture may be partly responsible for fewer Ca2+ sparks and a decrease in global Ca2+ release.

Background: The paper aimed to reveal the role and possible regulatory mechanisms of Wnt5 gene and its non-canonical Beta-catenin signaling pathway in osteogenic differentiation and bone repairment of hMSCs under infection environment. Material and Methods : CCK-8 method was employed to detect the effect of SPA on the proliferation of different hMSCs cells. The histological staining of alizarin red was used to observe the differentiation of cells into osteoblasts. The semi-quantitative detection of osteoblast alkaline phosphatase (ALP) was performed. Fluo-2 assay was employed to measure the intracellular calcium concentration of different hMSCs before and after osteogenic differentiation stimulated by SPA. RT-PCR was applied to detect the specific gene fluorescence of osteoblasts differentiation. Western Blotting was employed to determine the differentiation of hMSCs into osteoblasts. Results: Different hMSCs were able to proliferate in vitro under the stimulation of 100 ng/mL SPA. CCK-8 kit was used to detect lentivirus-infected hMSCs and non-transduced hMSCs in vitro. The results showed that all the cells gradually proliferate with time and an increasing rate. The hMSCs treated with different Wnt5 genes were stained with alizarin red after osteogenic differentiation, and the activities of AMSCs ALP increased. The intracellular free calcium concentration increased with the induction time and peaked in the Wnt5 overexpression group. PCR results showed that ALP, Runx-2, Collagen I, OC, Osterix and BSP genes were all increased in contrast with the non-induction Control group. Conclusion: Under the inflammatory condition of SPA virulence factors, non-canonical Wnt signaling pathway is associated with the classical Wnt/ β-catenin signaling pathway in the process of osteogenic differentiation of hMSCs in vitro, which has a certain synergistic effect.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that lead to the degeneration and death of motor neurons (MNs). Wnt signaling participates in multiple neurodegenerative processes. Wnt5a is a critical signaling molecule in Wnt signaling pathway. Intracellular Ca2+ homeostasis is disrupted in a number of neurodegenerative diseases, and Ca2+/Calmodulin dependent protein kinase II (CaMKII) involved in various diseases associated with Ca2+ signaling abnormalities. Here, we found that Wnt5a modulated MNs degeneration during ALS. CaMKII is a key effector of signals derived from Wnt/Ca2+ signaling pathway. Its main subtypes CaMKII-α and CaMKII-β were down-regulated in the spinal cord of SOD1G93A transgenic mice (ALS mice) and SOD1 mutant motor neuron like hybrid (NSC-34) cells. In addition, results showed that CaMKII-α and CaMKII-β were positively regulated by Wnt5a in vitro. Using specific CaMKII inhibitor and activator, we found that Wnt5a promoted NSC-34 cells viability and proliferation, inhibited cells apoptosis and promoted cells neurite outgrowth via the Wnt/Ca2+ signaling pathway. These results indicate that Wnt5a confers neuroprotection by promoting neuronal proliferation, inhibiting cell apoptosis and promoting neurite growth through Wnt/Ca2+ signaling pathway. Therefore, targeting Wnt5a can be an effective strategy to treat ALS.

Background Uterine serous cancer (USC) is the most common non-endometrioid subtype of uterine cancer, and is also the most aggressive. Most patients will die of progressively chemotherapy-resistant disease, and the development of new therapies that can target USC remains a major unmet clinical need. This study sought to determine the molecular mechanism by which a novel unfavorable prognostic biomarker RYR1 identified in advanced USC confers their malignant phenotypes, and demonstrated the efficacy of targeting RYR1 by repositioned FDA-approved compounds in USC treatment. Methods TCGA USC dataset was analyzed to identify top genes that are associated with patient survival and can be targeted by FDA-approved compounds. The top gene RYR1 was selected and the functional role of RYR1 in USC progression was determined by silencing and over-expressing RYR1 in USC cells in vitro and in vivo. The molecular mechanism and signaling networks associated with the functional role of RYR1 in USC progression were determined by reverse phase protein arrays (RPPA), Western blot, and transcriptomic profiling analyses. The efficacy of the repositioned compound dantrolene on USC progression was determined using both in vitro and in vivo models. Results High expression level of ryanodine receptor 1 (RYR1) in the tumors is associated with shortened overall survival. Inhibition of RYR1 suppressed proliferation, migration and enhanced apoptosis through the Ca2+-dependent AKT/CREB/PGC-1α and AKT/HK1/2 signaling pathways, which modulate mitochondrial bioenergetics properties, including oxidative phosphorylation, ATP production, mitochondrial membrane potential, ROS production and TCA metabolites, and glycolytic activities in USC cells. Repositioned compound dantrolene suppressed USC progression in both in vitro and mouse models. Conclusions These findings provide insight into the mechanism by which RYR1 modulates the malignant phenotypes of USC and could aid in the development of dantrolene as a repurposed therapeutic agent for the treatment of USC to improve patient survival.

Cancer is an aging-associated disease and caused by genomic instability that is driven by the accumulation of mutations and epimutations in the aging process. Although Ca2+ signaling, reactive oxygen species (ROS) accumulation, DNA damage response (DDR) and senescence inflammation response (SIR) are processed during genomic instability, the underlying mechanism for the cause of genomic instability and cancer development is still poorly understood and needs to be investigated. Nociceptive transient receptor potential (TRP) channels, which firstly respond to environmental stimuli, such as microbes, chemicals or physical injuries, potentiate regulation of the aging process by Ca2+ signaling. In this review, the authors provide an explanation of the dual role of nociceptive TRP channels in regulating cancer progression, initiating cancer progression by aging-induced genomic instability, and promoting malignancy by epigenetic regulation. Thus, therapeutically targeting nociceptive TRP channels seems to be a novel strategy for treating cancers.

Mutations in WFS1 (which encodes Wolframin, WFS1) and CISD2 (which encodes CDGSH iron sulfur domain 2) result in Wolfram syndrome (WS), a rare genetic disorder that starts with juvenile diabetes an...

SUMMARYIn mammalian sperm cells, regulation of spatiotemporal Ca2+ signaling relies on the quadrilinear Ca2+ signaling nanodomains in the flagellar membrane. The sperm-specific, multi-subunit CatSper Ca2+ channel, which is crucial for sperm hyperactivated motility and male fertility, organizes the nanodomains. Here, we report CatSperτ, the C2cd6-encoded membrane-associating C2 domain protein, can independently migrate to the flagella and serve as a major targeting component of the CatSper channel complex. CatSperτ loss-of-function in mice demonstrates that it is essential for sperm hyperactivated motility and male fertility. CatSperτ targets the CatSper channel into the quadrilinear nanodomains in the flagella of developing spermatids, whereas it is dispensable for functional channel assembly. CatSperτ interacts with ciliary trafficking machinery in a C2-dependent manner. These findings provide insights into the CatSper channel trafficking to the Ca2+ signaling nanodomains and the shared molecular mechanisms of ciliary and flagellar membrane targeting.HighlightsCatSperτ encoded by C2cd6 is a C2 membrane-associating domain containing proteinCatSperτ loss-of-function impairs sperm hyperactivation and male fertilityCatSperτ adopts ciliary trafficking machineries for flagellar targeting via C2 domainCatSperτ targets the CatSper channel into nanodomains of developing sperm flagella

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text In metaphase II arrested mammalian oocytes (eggs) and cleavage stage embryos the mitochondria are responsible for nearly all ATP production because glycolysis is inactivated. Luciferase assays show that ATP levels in eggs are strictly dependent upon pyruvate and fatty acid oxidation. The level of ATP in eggs appears to be maximal in conventional medium because the addition of extra mitochondrial substrates to eggs does not increase cytosolic ATP. The only clear elevation of ATP is seen at fertilization and is associated with sperm induced Ca2+ oscillations. Our recent findings suggest that the level of ATP modulates events at fertilization. At fertilization, the egg is activated by sperm derived PLCzeta which triggers a series of Ca2+ oscillations, with each Ca2+ release event causes by inositol trisphosphate (InsP3). Previous studies have shown that mouse eggs are more sensitive to PLCzeta, and generate higher frequency Ca2+ oscillations, than human eggs. Mouse eggs also generate Ca2+ oscillations and activate in response to Sr2+ that directly stimulates InsP3 receptors. In contrast, human eggs that contain the same type of InsP3 receptors do not generate Ca2+ oscillations in response to Sr2+. The difference in sensitivity of Ca2+ release between species can be explained by the fact that mouse eggs are about ten times more sensitive to InsP3 than human eggs. The reason for this difference appears to be due to ATP. The ATP level in unfertilized mouse eggs is about twice that in human eggs. Furthermore, the ability of mouse eggs to Sr2+ medium can be abolished by removing the mitochondrial substrate pyruvate, which reduces the ATP level. Adding back pyruvate to such eggs restores ATP levels promotes Sr2+ induced Ca2+ levels in mouse eggs. These data suggest that the level of ATP, possibly as ATP4-, modulates the sensitivity of the InsP3 receptor and the ability of eggs to generate Ca2+ oscillations. The level of cytosolic ATP may represent a significant ‘egg factor’ in determining the success of fertilization in humans. Enhancing mitochondrial ATP production could be useful in improving activation and embryo development after fertilization, or after artificial egg activation. References: Dumollard et al. (2009) Seminar in Cell and Developmental Biology 20, 346-353 Campbell and Swann (2006) Developmental Biology 298, 225-233 Storey et al. (2021) Molecular Human Reproduction 27, gaaa086

Oxysterol-binding protein–related protein (ORP) 4L acts as a scaffold protein assembling CD3-e, G-αq/11, and PLC-β3 into a complex at the plasma membrane that mediates inositol (1,4,5)-trisphosphat...

Canonical members of the TRP superfamily of ion channels have long been recognized as key elements of Ca2+ handling in a plethora of cell types. The emerging role of TRPC channels in human physiopathology has generated considerable interest in their pharmacological targeting, which requires detailed understanding of their molecular function. Although consent has been reached that receptor-phospholipase C (PLC) pathways and generation of lipid mediators constitute the prominent upstream signaling process that governs channel activity, multimodal sensing features of TRPC complexes have been demonstrated repeatedly. Downstream signaling by TRPC channels is similarly complex and involves the generation of local and global cellular Ca2+ rises, which are well-defined in space and time to govern specific cellular functions. These TRPC-mediated Ca2+ signals rely in part on Ca2+ permeation through the channels, but are essentially complemented by secondary mechanisms such as Ca2+ mobilization from storage sites and Na+/Ca2+ exchange, which involve coordinated interaction with signaling partners. Consequently, the control of cell functions by TRPC molecules is critically determined by dynamic assembly and subcellular targeting of the TRPC complexes. The very recent availability of high-resolution structure information on TRPC channel complexes has paved the way towards a comprehensive understanding of signal transduction by TRPC channels. Here, we summarize current concepts of cation permeation in TRPC complexes, TRPC-mediated shaping of cellular Ca2+ signals and the associated control of specific cell functions.

Ca2+ is essential for cellular functions. The ability of this ion to act as an intracellular signal, common to almost all eukaryotic cells, allows it to orchestrate an enormous repertoire of cellular activities, from initiating egg development after fertilization to synaptic transmission between neurons.

Physiological signaling by reactive oxygen species (ROS) and their pathophysiological role in cell death are well recognized. This review focuses on two ROS targets that are key to local Ca(2+) signaling at the ER/mitochondrial interface - notably, inositol trisphosphate receptors (IP(3)Rs) and the mitochondrial calcium uniporter (MCU). Both transport systems are central to molecular mechanisms in cell survival and death. Methods for the measurement of the redox state of these proteins and for the detection of ROS nanodomains are described. Recent results on the redox regulation of these proteins are reviewed.

Aims The effects of acute (100 s) hypoxia and/or acidosis on Ca2+ signaling parameters of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are explored here for the first time. Methods and results 1) hiPSC-CMs express two cell populations: rapidly-inactivating ICa myocytes (τi Conclusions Variability in ICa inactivation kinetics rather than cellular ancestry seems to underlie the action potential morphology differences generally attributed to mixed atrial and ventricular cell populations in hiPSC-CMs cultures. The differential hypoxic regulation of Ca2+-signaling in the two-cell types arises from differential Ca2+-dependent inactivation of the Ca2+-channel caused by proximity of Ca2+-release stores to the Ca2+ channels.

Summary: Leukemia stem cells (LSCs) are a rare subpopulation of abnormal hematopoietic stem cells (HSCs) that propagates leukemia and are responsible for the high frequency of relapse in therapies. Detailed insights into LSCs’ survival will facilitate the identification of targets for therapeutic approaches. Here, we develop an inhibitor, LYZ-81, which targets ORP4L with high affinity and specificity and selectively eradicates LCSs in vitro and in vivo. ORP4L is expressed in LSCs but not in normal HSCs and is essential for LSC bioenergetics and survival. It extracts PIP2 from the plasma membrane and presents it to PLCβ3, enabling IP3 generation and subsequent Ca2+-dependent bioenergetics. LYZ-81 binds ORP4L competitively with PIP2 and blocks PIP2 hydrolysis, resulting in defective Ca2+ signaling. The results provide evidence that LSCs can be eradicated through the inhibition of ORP4L by LYZ-81, which may serve as a starting point of drug development for the elimination of LSCs to eventually cure leukemia. : Zhong et al. report that abnormal expression of ORP4L is essential for leukemia stem cell survival; it enables IP3 generation by extracting and presenting PIP2 from the plasma membrane to PLCβ3 for hydrolysis. The compound LYZ-81, which blocks this process via targeting ORP4L, selectively eradicates leukemia stem cells. Keywords: leukemia stem cell, OSBP-related protein 4L, PIP2 hydrolysis, Ca2+ signaling, energy metabolism, therapy

Aims: Glucagon-like peptide-1 (GLP-1) increases intracellular Ca2+ concentrations, resulting in insulin secretion from pancreatic β-cells through the sequential production of Ca2+ mobilizi...

Terfenadine, an antihistamine used for the treatment of allergic conditions, affected Ca2+-related physiological responses in various models. However, the effect of terfenadine on cytosolic free Ca...

For the function and survival of neurons in the central nervous system, it is indispensable that the intracellular Ca2+ dynamics are properly controlled. The endoplasmic reticulum (ER), a major intracellular Ca2+ store, plays an important role in the control of the intracellular Ca2+ dynamics in neurons through the supply and uptake of Ca2+. It has been suggested that the disruption of ER Ca2+ signaling is involved in neuronal cell death in various pathological conditions. Therefore, the disruption of ER Ca2+ signaling has attracted attention as a novel mechanism for neurodegenerative diseases including Alzheimer's disease. In this review, we introduce the latest findings including our research on the relationship between the disruption of ER Ca2+ signaling and neuronal cell death. In addition, we will introduce recent developments on the technology for visualizing intraluminal Ca2+ dynamics within the ER, which is indispensable for promoting research in this field.

Transient receptor potential cation channel subfamily V member 4 (TRPV4)-mediated Ca2+signaling induces early activation of small/intermediate Ca2+-activated K+channels, SK3 (KCNN3) and IK1 (KCNN4), which leads to membrane hyperpolarization and enhanced Ca2+influx, which is critical for subsequent activation of the large conductance Ca2+-activated K+channel BK (KCNMA1) and K+secretion in kidney cortical collecting duct (CCD) cells. The focus of the present study was to determine if such coordinated hierarchical/sequential activation of these channels in CCD was orchestrated within caveolae, a known microcompartment underlying selective Ca2+-signaling events in other cells. In K+-secreting mouse principal cell (PC) line, mCCDcl1 cells, knockdown of caveolae caveolin-1 (CAV-1) depressed TRPV4-mediated Ca2+signaling and activation of SK3, intermediate conductance channel (IK1), and BK. Immunofluorescence colocalization analysis and coimmunoprecipitation assays demonstrated direct coupling of TRPV4 with each of the KCa channels in both mCCDcl1 and whole mouse kidney homogenates. Likewise, extending this analysis to CAV-1 demonstrates colocalization and direct coupling of CAV-1 with TRPV4, SK3, IK1, and BK, providing strong support for coupling of the channels in caveolae microdomains. Furthermore, differential expression of CAV-1 along the CCD was apparent where CAV-1 was strongly expressed within and along the cell borders of kidney PCs and intercalated cells (ICs), although significantly less in ICs. It is concluded that caveolae provide a key microdomain in PCs and ICs for coupling of TRPV4 with SK3, IK1, and BK that directly contributes to TRPV4-mediated Ca2+signaling in these domains leading to rapid and sequential coupling of TRPV4-SK3/IK1-BK that may play a central role in mediating Ca2+-dependent regulation of BK and K+secretion.

Mitochondrial Ca(2+) regulation is crucial for bioenergetics and cellular signaling. The mechanisms controlling mitochondrial calcium homeostasis have been recently unraveled with the discovery of mitochondrial inner membrane proteins that regulate mitochondrial Ca(2+) uptake and extrusion. Mitochondrial Ca(2+) uptake depends on a large complex of proteins centered around the Ca(2+) channel protein, mitochondrial Ca(2+) uniporter (MCU) in close interactions with several regulatory subunits (MCUb, EMRE, MICU1, MICU2). Mitochondrial Ca(2+) extrusion is mainly mediated by the mitochondrial Na(+)/Ca(2+)/Li(+) exchanger (NCLX). Here, we review the major players of mitochondrial Ca(2+) homeostasis and their physiological functions.

Intracellular Ca2+ ions that are thought to be one of the most important second messengers for cellular signaling, have a substantial diversity of roles in regulating a plethora of fundamental cellular physiology such as gene expression, cell division, cell motility and apoptosis. It has been suggestive of the Ca2+ signaling-dependent cellular processes to be tightly regulated by the numerous types of Ca2+ channels, pumps, exchangers and sensing receptors. Consequently, dysregulated Ca2+ homeostasis leads to a series of events connected to elevated malignant phenotypes including uncontrolled proliferation, migration, invasion and metastasis, all of which are frequently observed in advanced stage lung cancer cells. The incidence of bone metastasis in patients with advanced stage lung cancer is estimated in a range of 30% to 40%, bringing about a significant negative impact on both morbidity and survival. This review dissects and summarizes the important roles of Ca2+ signaling transduction in contributing to lung cancer progression, and address the question: if and how Ca2+ signaling might have been engaged in metastatic lung cancer with bone metastasis, thereby potentially providing the multifaceted and promising solutions for therapeutic intervention.

Circadian rhythms are based on biochemical oscillations generated by clock genes/proteins, which independently evolved in animals, fungi, plants, and cyanobacteria. Temperature compensation of the oscillation speed is a common feature of the circadian clocks, but the evolutionary-conserved mechanism has been unclear. Here, we show that Na+/Ca2+ exchanger (NCX) mediates cold-responsive Ca2+ signaling important for the temperature-compensated oscillation in mammalian cells. In response to temperature decrease, NCX elevates intracellular Ca2+, which activates Ca2+/calmodulin-dependent protein kinase II and accelerates transcriptional oscillations of clock genes. The cold-responsive Ca2+ signaling is conserved among mice, Drosophila, and Arabidopsis. The mammalian cellular rhythms and Drosophila behavioral rhythms were severely attenuated by NCX inhibition, indicating essential roles of NCX in both temperature compensation and autonomous oscillation. NCX also contributes to the temperature-compensated transcriptional rhythms in cyanobacterial clock. Our results suggest that NCX-mediated Ca2+ signaling is a common mechanism underlying temperature-compensated circadian rhythms both in eukaryotes and prokaryotes.

Simple Summary Intracellular Ca2+ signaling is a critical factor in breast cancer metastasis. In the proliferation stage, increases in intracellular Ca2+ concentration through voltage-dependent Ca2+ channels, P2Y2 channels, transient receptor potential (TRP) channels, store-operated Ca2+ channels (SOCCs), and IP3 receptors and a decrease in intracellular Ca2+ concentration through plasma membrane Ca2+ ATPases and secretory pathway Ca2+ ATPases (SPCA) activate breast cancer cell proliferation. TRPM7, SOCC, inositol trisphosphate receptor (IP3R), ryanodine receptor (RyR), and sarco-/endo-plasmic reticulum Ca2+-ATPase (SERCA) increase the expression of epithelial-to-mesenchymal transition (EMT)-related proteins; meanwhile, SPCA and the Na+/Ca2+ exchanger (NCX) control the activation of EMT-related proteins. Increased Ca2+ through TRPC1, TRPM7/8, P2X7, and SOCC enhances breast cancer cell migration. The stromal interaction molecule (STIM)-Orai complex, P2X7, and Ca2+ sensing receptors are involved in invadopodia. Various pharmacological agents for Ca2+ channels have been proposed against breast cancer and have provided potential strategies for treating metastatic processes. Abstract Metastatic features of breast cancer in the brain are considered a common pathology in female patients with late-stage breast cancer. Ca2+ signaling and the overexpression pattern of Ca2+ channels have been regarded as oncogenic markers of breast cancer. In other words, breast tumor development can be mediated by inhibiting Ca2+ channels. Although the therapeutic potential of inhibiting Ca2+ channels against breast cancer has been demonstrated, the relationship between breast cancer metastasis and Ca2+ channels is not yet understood. Thus, we focused on the metastatic features of breast cancer and summarized the basic mechanisms of Ca2+-related proteins and channels during the stages of metastatic breast cancer by evaluating Ca2+ signaling. In particular, we highlighted the metastasis of breast tumors to the brain. Thus, modulating Ca2+ channels with Ca2+ channel inhibitors and combined applications will advance treatment strategies for breast cancer metastasis to the brain.

A Correction to this paper has been published: https://doi.org/10.1038/s42003-021-01717-x

Inositol trisphosphate (IP3) is a Ca2+-mobilizing second messenger shown to modulate atrial muscle contraction, and is thought to contribute to atrial fibrillation. Cellular pathways underlying IP3 actions in cardiac tissue remain poorly understood, and the work presented here addresses the question whether IP3-mediated Ca2+ release from the sarcoplasmic reticulum is linked to adenylyl cyclase activity including Ca2+-stimulated adenylyl cyclases (AC1 and AC8) that are selectively expressed in atria and sino-atrial node (SAN). Immunocytochemistry in guinea pig atrial myocytes identified co-localization of type 2 IP3Rs with AC8, while AC1 was located in close vicinity. Intracellular photorelease of IP3 by UV light significantly enhanced the amplitude of the Ca2+ transient (CaT) evoked by electrical stimulation of atrial myocytes (31 ± 6 % increase 60 s post photorelease, n=16). The increase in CaT amplitude was abolished by inhibitors of adenylyl cyclases (MDL-12,330) or protein kinase A (H89), showing that cAMP signaling is required for this effect of photoreleased IP3. In mouse spontaneously beating right atrial preparations, phenylephrine, an α-adrenoceptor agonist with effects that depend on IP3 mediated Ca2+ release, increased the maximum beating rate by 14.7 ± 0.5 %, n=10. This effect was substantially reduced by 2.5 µmol/L 2-APB and abolished by a low dose of MDL-12,330, observations which are again consistent with a functional interaction between IP3 and cAMP signaling involving Ca2+ stimulation of adenylyl cyclases in the SAN pacemaker. Understanding the interaction between IP3 receptor pathways and Ca2+-stimulated adenylyl cyclases provides important insights concerning acute mechanisms for initiation of atrial arrhythmias.

Recent achievements in indicator optimization and imaging techniques promote the advancement of functional imaging to decipher complex signaling processes in living cells, such as Ca2+ activity patterns. Astrocytes are important regulators of the brain network and well known for their highly complex morphology and spontaneous Ca2+ activity. However, the astrocyte community is lacking standardized methods to analyze and interpret Ca2+ activity recordings, hindering global comparisons. Here, we present a biophysically-based analytical concept for deciphering the complex spatio-temporal changes of Ca2+ biosensor fluorescence for understanding the underlying signaling mechanisms. We developed a pixel-based multi-threshold event detection (MTED) analysis of multidimensional data, which accounts for signal strength as an additional signaling dimension and provides the experimenter with a comprehensive toolbox for a differentiated and in-depth characterization of fluorescence signals. MTED was validated by analyzing astrocytic Ca2+ activity across Ca2+ indicators, imaging setups, and model systems from primary cell culture to awake, head-fixed mice. We identified extended Ca2+ activity at 25°C compared to 37°C physiological body temperature and dissected how neuronal activity shapes long-lasting astrocytic Ca2+ activity. Our MTED strategy, as a parameter-free approach, is easily transferrable to other fluorescent indicators and biosensors and embraces the additional dimensionality of signaling activity strength. It will also advance the definition of standardized procedures and parameters to improve comparability of research data and reports.

Mineral nutrition plays a significant role in plant development. Initial studies focused on calcium (Ca2+) as an important plant nutrient. Later researchers identified that Ca2+ was part of a vast signaling network that was developed due to biochemical properties of Ca2+. Plants have evolved separate Ca2+ homeostasis and Ca2+ binding proteins than animals. However, these differences do not hinder the growth and development in plants, and the plant Ca2+ signaling toolkit is optimized for catering to its needs.

Background Dilated cardiomyopathy (DCM) is among the most common causes leading to end-stage heart failure with reduced ejection fraction (HF-rEF) in adult and pediatric patients. Despite similar phenotypes characterized as systolic dysfunction and eccentric ventricular dilation, pediatric DCM are biologically distinct entities with age- and development-specific features in the heart. Though underlying mechanisms may vary between the two populations, it's largely unexplored with few studies conducted to date. Purpose HF-rEF typically results from impaired myocardial contractility, triggered by defective cellular Ca2+ handling and cytoskeletal remodeling. Hence, we aim to integrate clinical profile and experimental data from human explanted hearts: 1) to unravel the age-dependent disparate Ca2+ signaling pathways; and 2) to identify pediatric-specific HF signatures or potential cures for precision managements. Methods Non-ischemic failing hearts (n=6 adult and n=6 pediatric) were procured immediately after excision via Human Explanted Heart Program. Age-matched adult non-failing control hearts (NFC, n=6) were obtained from deceased donors without cardiovascular history, while pediatric NFC (n=6) were collected from children with congenital heart defects but no primary myocardial dysfunction constituting relatively reasonable controls. Myocardial metabolic and oxidative profile were evaluated spectrophotometrically, and tissue remodeling was assessed immunohistochemically. Global proteomics and phosphoproteomics were performed on a Q-Exactive mass spectrometer, followed by network biology pathway analyses. Expression of screened proteins and kinases was validated by gel electrophoresis. Apoptosis and cellular growth signaling pathways were also incorporated into analysis. Results Both HF groups had remarkably lower LVEF (26.6±10.7% in pediatric vs. 26.5±9.1% in adult DCM) while compared to the NFC (both ≥60%) respectively. Histologically, adult-DCM demonstrated significantly worse fibrosis than pediatric-DCM (p Conclusions Pediatric DCM exhibited less adverse remodeling partially mediated by divergent Ca2+ handling and downstream signaling pathways, illustrating the fundamental differences between adult and pediatric DCM. Our findings may provide a scientific basis for the development of specific therapies for pediatric DCM. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Canadian Institutes for Health Research (CIHR); Heart & Stroke Foundation (HSF)

Interstitial cells of Cajal (ICC) generate pacemaker activity responsible for phasic contractions in colonic segmentation and peristalsis. ICC along the submucosal border (ICC-SM) contributing to mixing and more complex patterns of colonic motility. We show the complex patterns of Ca2+signaling in ICC-SM and the relationship between ICC-SM Ca2+transients and activation of SMCs using optogenetic tools. ICC-SM displayed rhythmic firing of Ca2+transients ∼15 cpm and paced adjacent SMCs. The majority of spontaneous activity occurred in regular Ca2+ transients clusters (CTCs) that propagated through the network. CTCs were organized and dependent upon Ca2+entry through voltage-dependent Ca2+conductances, L- and T-type Ca2+channels. Removal of Ca2+from the external solution abolished CTCs. Ca2+release mechanisms reduced the duration and amplitude of Ca2+transients but did not block CTCs. These data reveal how colonic pacemaker ICC-SM exhibit complex Ca2+firing patterns and drive smooth muscle activity and overall colonic contractions.SynopsisHow Ca2+signaling in colonic submucosal pacemaker cells couples to smooth muscle responses is unknown. This study shows how ICC modulate colonic motility via complex Ca2+signaling and defines Ca2+transients’ sources using optogenetic techniques.

Understanding ER-to-mitochondrial Ca 2+ flux may provide ways to correct mitochondrial dysfunction in various diseases.

Cells of the oligodendrocyte lineage express a wide range of Ca2+ channels and receptors that regulate oligodendrocyte progenitor cell (OPC) and oligodendrocyte formation and function. Here we define those key channels and receptors that regulate Ca2+ signaling and OPC development and myelination. We then discuss how the regulation of intracellular Ca2+ in turn affects OPC and oligodendrocyte biology in the healthy nervous system and under pathological conditions. Activation of Ca2+ channels and receptors in OPCs and oligodendrocytes by neurotransmitters converges on regulating intracellular Ca2+, making Ca2+ signaling a central candidate mediator of activity-driven myelination. Indeed, recent evidence indicates that localized changes in Ca2+ in oligodendrocytes can regulate the formation and remodeling of myelin sheaths and perhaps additional functions of oligodendrocytes and OPCs. Thus, decoding how OPCs and myelinating oligodendrocytes integrate and process Ca2+ signals will be important to fully understand central nervous system formation, health, and function.

Navigation of sperm in fluid flow, called rheotaxis, provides long‐range guidance in the mammalian oviduct. The rotation of sperm around their longitudinal axis (rolling) promotes rheotaxis. Whether sperm rolling and rheotaxis require calcium (Ca2+) influx via the sperm‐specific Ca2+ channel CatSper, or rather represent passive biomechanical and hydrodynamic processes, has remained controversial. Here, we study the swimming behavior of sperm from healthy donors and from infertile patients that lack functional CatSper channels, using dark‐field microscopy, optical tweezers, and microfluidics. We demonstrate that rolling and rheotaxis persist in CatSper‐deficient human sperm. Furthermore, human sperm undergo rolling and rheotaxis even when Ca2+ influx is prevented. Finally, we show that rolling and rheotaxis also persist in mouse sperm deficient in both CatSper and flagellar Ca2+‐signaling domains. Our results strongly support the concept that passive biomechanical and hydrodynamic processes enable sperm rolling and rheotaxis, rather than calcium signaling mediated by CatSper or other mechanisms controlling transmembrane Ca2+ flux., Live‐cell imaging unexpectedly reveals that Ca2+ influx via the CatSper channel is dispensable for rheotactic steering of sperm.

Pacemaker action potentials emerge from the sinoatrial node (SAN) and rapidly propagate through the atria to the AV node via preferential conduction pathways, including one associated with the coronary sinus. However, few distinguishing features of these tracts are known. Identifying specific molecular markers to distinguish among these conduction pathways will have important implications for understanding atrial conduction and atrial arrhythmogenesis. Using a Stim1 reporter mouse, we discovered stromal interaction molecule 1 (STIM1)-expressing coronary sinus cardiomyocytes (CSC)s in a tract from the SAN to the coronary sinus. Our studies here establish that STIM1 is a molecular marker of CSCs and we propose a role for STIM1-CSCs in interatrial conduction. Deletion of Stim1 from the CSCs slowed interatrial conduction and increased susceptibility to atrial arrhythmias. Store-operated Ca(2+) currents (I(soc)) in response to Ca(2+) store depletion were markedly reduced in CSCs and their action potentials showed electrical remodeling. Our studies identify STIM1 as a molecular marker for a coronary sinus interatrial conduction pathway. We propose a role for SOCE in Ca(2+) signaling of CSCs and implicate STIM1 in atrial arrhythmogenesis.

Inter-organelle contacts are emerging as crucial hubs in cell signaling. Among these, the long known ER-mitochondria tethers can be considered a reference model for these communication structures. Here, signal crosstalk at the interface of ER and mitochondria occurs thanks to lipids exchange and Ca2+ flux across these specialized structures. How this signal is modulated depends on the compositions of the proteins that are engaged at these areas of membrane proximity. These proteins modulate the activity of Ca2+ channels and regulate contact structure remodeling in response to cellular cues. In this review, we highlight the molecular aspects of the ER-mitochondria contacts and how they impact on Ca2+ signaling and thus on cell physiology.

Vascular endothelial cells (ECs) sense and transduce hemodynamic shear stress into intracellular biochemical signals, and Ca2+signaling plays a critical role in this mechanotransduction, i.e., ECs release ATP in the caveolae in response to shear stress and, in turn, the released ATP activates P2 purinoceptors, which results in an influx into the cells of extracellular Ca2+. However, the mechanism by which the shear stress evokes ATP release remains unclear. Here, we demonstrated that cellular mitochondria play a critical role in this process. Cultured human pulmonary artery ECs were exposed to controlled levels of shear stress in a flow-loading device, and changes in the mitochondrial ATP levels were examined by real-time imaging using a fluorescence resonance energy transfer-based ATP biosensor. Immediately upon exposure of the cells to flow, mitochondrial ATP levels increased, which was both reversible and dependent on the intensity of shear stress. Inhibitors of the mitochondrial electron transport chain and ATP synthase as well as knockdown of caveolin-1, a major structural protein of the caveolae, abolished the shear stress-induced mitochondrial ATP generation, resulting in the loss of ATP release and influx of Ca2+into the cells. These results suggest the novel role of mitochondria in transducing shear stress into ATP generation: ATP generation leads to ATP release in the caveolae, triggering purinergic Ca2+signaling. Thus, exposure of ECs to shear stress seems to activate mitochondrial ATP generation through caveola- or caveolin-1-mediated mechanisms.NEW & NOTEWORTHY The mechanism of how vascular endothelial cells sense shear stress generated by blood flow and transduce it into functional responses remains unclear. Real-time imaging of mitochondrial ATP demonstrated the novel role of endothelial mitochondria as mechanosignaling organelles that are able to transduce shear stress into ATP generation, triggering ATP release and purinoceptor-mediated Ca2+signaling within the cells.

The migration of endothelial cells (ECs) is closely associated with a Ca2+ -dependent protein, protein kinase Cα (PKCα). The disruption of intercellular adhesion by single-cell wounding has been shown to induce the directional translocation of PKCα. We hypothesized that this translocation of PKCα is induced by mechanical stress, such as unloading of intercellular tension, or by intercellular communication, such as gap junction-mediated and paracrine signaling. In the current study, we found that the disruption of intercellular adhesion induced the directional translocation of PKCα even when gap junction-mediated and paracrine signaling were inhibited. Conversely, it did not occur when the mechanosensitive channel was inhibited. In addition, the strain field of substrate attributable to the disruption of intercellular adhesion tended to be larger at the areas corresponding with PKCα translocation. Recently, we found that a direct mechanical stimulus induced the accumulation of PKCα at the stimulus area, involving Ca 2+ influx from extracellular space. These results indicated that the unloading of intercellular tension induced directional translocation of PKCα, which required Ca 2+ influx from extracellular space. The results of this study indicate the involvement of PKCα in the Ca 2+ signaling pathway in response to mechanical stress in ECs.

Background Recently, pathogenic alleles within ubiquitin N-recognin domain-containing E3 ligase 4 (UBR4) gene have been shown to be associated with Hirschsprung disease (HSCR). We determined the UBR4 expressions in Indonesian HSCR patients. Methods We analyzed the UBR4 expressions in the colons of HSCR patient and anorectal malformation (ARM) patient as control by real-time polymerase chain reaction (qPCR). Results Thirty-seven patients with non-syndromic HSCR and eighteen controls were involved in this study. qPCR revealed that the UBR4 expression was strongly decreased (0.77-fold) in the ganglionic group of patients with HSCR compared to the control group with ARM (ΔCT 2.43 ± 0.36 vs. 2.05 ± 0.69; p = 0.009), whereas the UBR4 expression was also significantly reduced (0.79-fold) in the aganglionic group of patients with HSCR compared to the control group with ARM (ΔCT 2.39 ± 0.46 vs. 2.05 ± 0.69; p = 0.044). However, the UBR4 expression change was not associated with gender (p = 0.35 and 0.80), nor with degree of aganglionosis both in ganglionic and aganglionic colons (p = 0.72 and 0.73), respectively. Conclusion Our study demonstrates that expression of UBR4 is decreased in both aganglionic and ganglionic colon of HSCR patients.

Physiological functions of type 3 ryanodine receptors (RyR3) in smooth muscle (SM) tissues are not well understood, in spite of their wide expression. However, the short isoform of RyR3 is known to be a dominant-negative variant (DN-RyR3), which may negatively regulate functions of both RyR2 and full-length (FL) RyR3 by forming hetero-tetramers. Here, functional roles of RyR3 in the regulation of Ca2+signaling in mesenteric artery SM cells (MASMCs) were examined using RyR3 homozygous knockout mice (RyR3−/−). Quantitative PCR analyses suggested that the predominant RyR3 subtype in MASMs from wild-type mice (RyR3+/+) was DN-RyR3. In single MASMCs freshly isolated from RyR3−/−, the EC50of caffeine to induce Ca2+release was lower than that in RyR3+/+myocytes. The amplitude and frequency of Ca2+sparks and spontaneous transient outward currents in MASMCs from RyR3−/−were all larger than those from RyR3+/+. Importantly, mRNA and functional expressions of voltage-dependent Ca2+channel and large-conductance Ca2+-activated K+(BK) channel in MASMCs from RyR3−/−were identical to those from RyR3+/+. However, in the presence of BK channel inhibitor, paxilline, the pressure rises induced by BayK8644 in MA vascular beds of RyR3−/−were significantly larger than in those of RyR3+/+. This indicates that the negative feedback effects of BK channel activity on intracellular Ca2+signaling was enhanced in RyR3−/−. Thus, RyR3, and, in fact, mainly DN-RyR3, via a complex with RyR2 suppresses Ca2+release and indirectly regulated membrane potential by reducing BK channel activity in MASMCs and presumably can affect the regulation of intrinsic vascular tone.

The aim of the present study was to examine whether transient receptor potential melastatin 2 (TRPM2) plays a role in muscle fiber-type transition during exercise. Mice were trained at a speed of 12 m/min at a slope of 0° for 60 min for 5 consecutive days/wk for 4 wk. Exhaustion tests were performed on the treadmill (the speed was set at 6 m/min at a slope of 0° and increased at a rate of 1 m/min every 6 min). Isolated primary skeletal muscle cells from TRPM2-knockout (KO) mice showed lower amplitudes of electrical stimuli (ES)-induced Ca2+ signals when compared with wild-type (WT) mice due to a defect in Ca2+ influx. Moreover, TRPM2-KO mice had a higher proportion of fast-twitch skeletal muscle fibers and a lower proportion of slow-twitch muscle fibers before exercise than WT mice. After exercise, the expression of slow-twitch skeletal muscle fibers was increased only in WT mice but not in TRPM2-KO mice. ES-induced nuclear translocation of the Ca2+-dependent transcription factor NFATc1 was significantly lower in TRPM2-KO mice than in WT mice. TRPM2-KO mice also showed decreased mitochondrial Ca2+ and membrane potential. Lactate levels were higher in the skeletal muscle cells of TRPM2-KO mice before and after ES compared with WT mice. Collectively, these data indicate that TRPM2-mediated Ca2+ signaling plays a critical role in the regulation of fiber-type switching and mitochondrial function in skeletal muscle. NEW & NOTEWORTHY TRPM2 has been shown to play an important role in a variety of cellular functions. However, the role of TRPM2 in skeletal muscle remains poorly understood. Here, we provide evidence that TRPM2-mediated Ca2+ signaling is required for training-induced improvement in skeletal muscle mitochondrial function and fiber type transition.