Search Results (10)

Cardiac fibrosis is a scarring event that occurs in the myocardium in response to multiple cardiovascular disorders, such as acute myocardial infarction (AMI), ischemic cardiomyopathy, dilated cardiomyopathy, hypertensive heart disease, inflammatory heart disease, diabetic cardiomyopathy, and aortic stenosis. Fibrotic remodeling is mainly sustained by the differentiation of fibroblasts into myofibroblasts, which synthesize and secrete most of the extracellular matrix (ECM) proteins. An increase in the intracellular Ca²⁺ concentration ([Ca²⁺]_i) in cardiac fibroblasts is emerging as a critical mediator of the fibrogenic signaling cascade. Herein, we review the mechanisms that may shape intracellular Ca²⁺ signals involved in fibroblast transdifferentiation into myofibroblasts. We focus our attention on the functional interplay between inositol-1,4,5-trisphosphate (InsP₃) receptors (InsP₃Rs) and store-operated Ca²⁺ entry (SOCE). In accordance with this, InsP₃Rs and SOCE drive the Ca²⁺ response elicited by G_q-protein coupled receptors (G_qPCRs) that promote fibrotic remodeling. Then, we describe the additional mechanisms that sustain extracellular Ca²⁺ entry, including receptor-operated Ca²⁺ entry (ROCE), P2X receptors, Transient Receptor Potential (TRP) channels, and Piezo1 channels. In parallel, we discuss the pharmacological manipulation of the Ca²⁺ handling machinery as a promising approach to mitigate or reverse fibrotic remodeling in cardiac disorders. Full article

Multiple inositol polyphosphate phosphatase (MINPP1) is an enigmatic enzyme that is responsible for the metabolism of inositol hexakisphosphate (InsP₆) and inositol 1,3,4,5,6 pentakisphosphate (Ins(1,3,4,5,6)P₅ in mammalian cells, despite being restricted to the confines of the ER. The reason for this compartmentalization is unclear. In our previous studies in the insulin-secreting HIT cell line, we expressed MINPP1 in the cytosol to artificially reduce the concentration of these higher inositol phosphates. Undocumented at the time, we noted cytosolic MINPP1 expression reduced cell growth. We were struck by the similarities in substrate preference between a number of different enzymes that are able to metabolize both inositol phosphates and lipids, notably IPMK and PTEN. MINPP1 was first characterized as a phosphatase that could remove the 3-phosphate from inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P₄)_. This molecule shares strong structural homology with the major product of the growth-promoting Phosphatidyl 3-kinase (PI3K), phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P₃) and PTEN can degrade both this lipid and Ins(1,3,4,5)P₄. Because of this similar substrate preference, we postulated that the cytosolic version of MINPP1 (cyt-MINPP1) may not only attack inositol polyphosphates but also PtdIns(3,4,5)P₃, a key signal in mitogenesis. Our experiments show that expression of cyt-MINPP1 in HIT cells lowers the concentration of PtdIns(3,4,5)P₃. We conclude this reflects a direct effect of MINPP1 upon the lipid because cyt-MINPP1 actively dephosphorylates synthetic, di(C4:0)PtdIns(3,4,5)P₃ in vitro. These data illustrate the importance of MINPP1′s confinement to the ER whereby important aspects of inositol phosphate metabolism and inositol lipid signaling can be separately regulated and give one important clarification for MINPP1′s ER seclusion. Full article

Store-operated Ca²⁺ entry (SOCE) is activated in response to the inositol-1,4,5-trisphosphate (InsP₃)-dependent depletion of the endoplasmic reticulum (ER) Ca²⁺ store and represents a ubiquitous mode of Ca²⁺ influx. In vascular endothelial cells, SOCE regulates a plethora of functions that maintain cardiovascular homeostasis, such as angiogenesis, vascular tone, vascular permeability, platelet aggregation, and monocyte adhesion. The molecular mechanisms responsible for SOCE activation in vascular endothelial cells have engendered a long-lasting controversy. Traditionally, it has been assumed that the endothelial SOCE is mediated by two distinct ion channel signalplexes, i.e., STIM1/Orai1 and STIM1/Transient Receptor Potential Canonical 1(TRPC1)/TRPC4. However, recent evidence has shown that Orai1 can assemble with TRPC1 and TRPC4 to form a non-selective cation channel with intermediate electrophysiological features. Herein, we aim at bringing order to the distinct mechanisms that mediate endothelial SOCE in the vascular tree from multiple species (e.g., human, mouse, rat, and bovine). We propose that three distinct currents can mediate SOCE in vascular endothelial cells: (1) the Ca²⁺-selective Ca²⁺-release activated Ca²⁺ current (I_CRAC), which is mediated by STIM1 and Orai1; (2) the store-operated non-selective current (I_SOC), which is mediated by STIM1, TRPC1, and TRPC4; and (3) the moderately Ca²⁺-selective, I_CRAC-like current, which is mediated by STIM1, TRPC1, TRPC4, and Orai1. Full article

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy whose chemoresistance and relapse persist as a problem despite significant advances in its chemotherapeutic treatments. Mitochondrial metabolism has emerged as an interesting therapeutic target given its essential role in maintaining bioenergetic and metabolic homeostasis. T-ALL cells are characterized by high levels of mitochondrial respiration, making them suitable for this type of intervention. Mitochondrial function is sustained by a constitutive transfer of calcium from the endoplasmic reticulum to mitochondria through the inositol 1,4,5-trisphosphate receptor (InsP3R), making T-ALL cells vulnerable to its inhibition. Here, we determine the bioenergetic profile of the T-ALL cell lines CCRF-CEM and Jurkat and evaluate their sensitivity to InsP3R inhibition with the specific inhibitor, Xestospongin B (XeB). Our results show that T-ALL cell lines exhibit higher mitochondrial respiration than non-malignant cells, which is blunted by the inhibition of the InsP3R. Prolonged treatment with XeB causes T-ALL cell death without affecting the normal counterpart. Moreover, the combination of XeB and glucocorticoids significantly enhanced cell death in the CCRF-CEM cells. The inhibition of InsP3R with XeB rises as a potential therapeutic alternative for the treatment of T-ALL. Full article

Experimental evidence highlights the involvement of the endoplasmic reticulum (ER)-mediated Ca²⁺ signals in modulating synaptic plasticity and spatial memory formation in the hippocampus. Ca²⁺ release from the ER mainly occurs through two classes of Ca²⁺ channels, inositol 1,4,5-trisphosphate receptors (InsP3Rs) and ryanodine receptors (RyRs). Calsequestrin (CASQ) and calreticulin (CR) are the most abundant Ca²⁺-binding proteins allowing ER Ca²⁺ storage. The hippocampus is one of the brain regions expressing CASQ, but its role in neuronal activity, plasticity, and the learning processes is poorly investigated. Here, we used knockout mice lacking both CASQ type-1 and type-2 isoforms (double (d)CASQ-null mice) to: a) evaluate in adulthood the neuronal electrophysiological properties and synaptic plasticity in the hippocampal Cornu Ammonis 1 (CA1) field and b) study the performance of knockout mice in spatial learning tasks. The ablation of CASQ increased the CA1 neuron excitability and improved the long-term potentiation (LTP) maintenance. Consistently, (d)CASQ-null mice performed significantly better than controls in the Morris Water Maze task, needing a shorter time to develop a spatial preference for the goal. The Ca²⁺ handling analysis in CA1 pyramidal cells showed a decrement of Ca²⁺ transient amplitude in (d)CASQ-null mouse neurons, which is consistent with a decrease in afterhyperpolarization improving LTP. Altogether, our findings suggest that CASQ deletion affects activity-dependent ER Ca²⁺ release, thus facilitating synaptic plasticity and spatial learning in post-natal development. Full article

Phosphoinositides (PI) form just a minor portion of the total phospholipid content in cells but are significantly involved in cancer development and progression. In several cancer types, phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P₃] and phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P₂] play significant roles in regulating survival, proliferation, invasion, and growth of cancer cells. Phosphoinositide-specific phospholipase C (PLC) catalyze the generation of the essential second messengers diacylglycerol (DAG) and inositol 1,4,5 trisphosphate (InsP₃) by hydrolyzing PtdIns(4,5)P₂. DAG and InsP₃ regulate Protein Kinase C (PKC) activation and the release of calcium ions (Ca²⁺) into the cytosol, respectively. This event leads to the control of several important biological processes implicated in cancer. PLCs have been extensively studied in cancer but their regulatory roles in the oncogenic process are not fully understood. This review aims to provide up-to-date knowledge on the involvement of PLCs in cancer. We focus specifically on PLCβ, PLCγ, PLCδ, and PLCε isoforms due to the numerous evidence of their involvement in various cancer types. Full article

Cardiomyocytes and myocardial sleeves dissociated from pulmonary veins (PVs) potentially generate ectopic automaticity in response to noradrenaline (NA), and thereby trigger atrial fibrillation. We developed a mathematical model of rat PV cardiomyocytes (PVC) based on experimental data that incorporates the microscopic framework of the local control theory of Ca²⁺ release from the sarcoplasmic reticulum (SR), which can generate rhythmic Ca²⁺ release (limit cycle revealed by the bifurcation analysis) when total Ca²⁺ within the cell increased. Ca²⁺ overload in SR increased resting Ca²⁺ efflux through the type II inositol 1,4,5-trisphosphate (IP₃) receptors (InsP₃R) as well as ryanodine receptors (RyRs), which finally triggered massive Ca²⁺ release through activation of RyRs via local Ca²⁺ accumulation in the vicinity of RyRs. The new PVC model exhibited a resting potential of −68 mV. Under NA effects, repetitive Ca²⁺ release from SR triggered spontaneous action potentials (APs) by evoking transient depolarizations (TDs) through Na⁺/Ca²⁺ exchanger (AP_TDs). Marked and variable latencies initiating AP_TDs could be explained by the time courses of the α1- and β1-adrenergic influence on the regulation of intracellular Ca²⁺ content and random occurrences of spontaneous TD activating the first AP_TD. Positive and negative feedback relations were clarified under AP_TD generation. Full article

Nicotinic acid adenine dinucleotide phosphate (NAADP) gates two-pore channels 1 and 2 (TPC1 and TPC2) to elicit endo-lysosomal (EL) Ca²⁺ release. NAADP-induced EL Ca²⁺ signals may be amplified by the endoplasmic reticulum (ER) through the Ca²⁺-induced Ca²⁺ release mechanism (CICR). Herein, we aimed at assessing for the first time the role of EL Ca²⁺ signaling in primary cultures of human metastatic colorectal carcinoma (mCRC) by exploiting Ca²⁺ imaging and molecular biology techniques. The lysosomotropic agent, Gly-Phe β-naphthylamide (GPN), and nigericin, which dissipates the ΔpH which drives Ca²⁺ refilling of acidic organelles, caused massive Ca²⁺ release in the presence of a functional inositol-1,4,5-trisphosphate (InsP₃)-sensitive ER Ca²⁺ store. Liposomal delivery of NAADP induced a transient Ca²⁺ release that was reduced by GPN and NED-19, a selective TPC antagonist. Pharmacological and genetic manipulations revealed that the Ca²⁺ response to NAADP was triggered by TPC1, the most expressed TPC isoform in mCRC cells, and required ER-embedded InsP₃ receptors. Finally, NED-19 and genetic silencing of TPC1 reduced fetal calf serum-induced Ca²⁺ signals, proliferation, and extracellular signal-regulated kinase and Akt phoshorylation in mCRC cells. These data demonstrate that NAADP-gated TPC1 could be regarded as a novel target for alternative therapies to treat mCRC. Full article

Membrane contact sites are regions of close apposition between two organelles, typically less than 30 nanometers apart, that facilitate transfer of biomolecules. The presence of contact sites has been demonstrated in yeast, plants, and mammalian cells. Here, we investigated the presence of such contact sites in Trypanosoma brucei. In mammalian cells, endoplasmic reticulum-mitochondria contact sites facilitate mitochondrial uptake of Ca²⁺ released by the ER-located inositol 1,4,5-trisphosphate receptor (InsP₃R). However, the InsP₃R in trypanosomes localizes to acidocalcisomes, which serve as major Ca²⁺ stores in these parasites. In this work, we have used super-resolution structured illumination microscopy and electron microscopy to identify membrane contact sites that exist between acidocalcisomes and mitochondria. Furthermore, we have confirmed the close association of these organelles using proximity ligation assays. Characterization of these contact sites may be a necessary starting point towards unraveling the role of Ca²⁺ in regulating trypanosome bioenergetics. Full article

One challenge in studying the second messenger inositol(1,4,5)-trisphosphate (InsP₃) is that it is present in very low amounts and increases only transiently in response to stimuli. To identify events downstream of InsP₃, we generated transgenic plants constitutively expressing the high specific activity, human phosphatidylinositol 4-phosphate 5-kinase Iα (HsPIPKIα). PIP5K is the enzyme that synthesizes phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P₂); this reaction is flux limiting in InsP₃ biosynthesis in plants. Plasma membranes from transgenic Arabidopsis expressing HsPIPKIα had 2–3 fold higher PIP5K specific activity, and basal InsP₃ levels in seedlings and leaves were >2-fold higher than wild type. Although there was no significant difference in photosynthetic electron transport, HsPIPKIα plants had significantly higher starch (2–4 fold) and 20% higher anthocyanin compared to controls. Starch content was higher both during the day and at the end of dark period. In addition, transcripts of genes involved in starch metabolism such as SEX1 (glucan water dikinase) and SEX4 (phosphoglucan phosphatase), DBE (debranching enzyme), MEX1 (maltose transporter), APL3 (ADP-glucose pyrophosphorylase) and glucose-6-phosphate transporter (Glc6PT) were up-regulated in the HsPIPKIα plants. Our results reveal that increasing the phosphoinositide (PI) pathway affects chloroplast carbon metabolism and suggest that InsP₃ is one component of an inter-organelle signaling network regulating chloroplast metabolism. Full article