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Lysosomes and the endoplasmic reticulum (ER) are Ca2+ stores mobilized by the second messengers NAADP and IP3, respectively. Here, we establish Ca2+ signals between the two sources as fundamental building blocks that couple local release to global changes in Ca2+. Cell-wide Ca2+ signals evoked by activation of endogenous NAADP-sensitive channels on lysosomes comprise both local and global components and exhibit a major dependence on ER Ca2+ despite their lysosomal origin. Knockout of ER IP3 receptor channels delays these signals, whereas expression of lysosomal TPC2 channels accelerates them. High-resolution Ca2+ imaging reveals elementary events upon TPC2 opening and signals coupled to IP3 receptors. Biasing TPC2 activation to a Ca2+-permeable state sensitizes local Ca2+ signals to IP3. This increases the potency of a physiological agonist to evoke global Ca2+ signals and activate a downstream target. Our data provide a conceptual framework to understand how Ca2+ release from physically separated stores is coordinated.

Ca2+ signalling plays an essential role in T cell activation, which is a key step to start an adaptive immune response. During the transition from a quiescent to a fully activated state, Ca2+ microdomains of reduced spatial and temporal extents develop in the junctions between the plasma membrane and the endoplasmic reticulum (ER). These microdomains rely on Ca2+ entry from the extracellular medium, via the ORAI1/STIM1/STIM2 system that mediates store operated Ca2+ entry Store operated calcium entry. The mechanism leading to local store depletion and subsequent Ca2+ entry depends on the activation state of the cells. The initial, smaller microdomains are triggered by D-myo-inositol 1,4,5-trisphosphate (IP3) signalling in response to T cell adhesion. T cell receptor (TCR)/CD3 stimulation then initiates nicotinic acid adenine dinucleotide phosphate signalling, which activates ryanodine receptors (RYR). We have recently developed a mathematical model to elucidate the spatiotemporal Ca2+ dynamics of the microdomains triggered by IP3 signalling in response to T cell adhesion (Gil et al. 2021). This reaction-diffusion model describes the evolution of the cytosolic and endoplasmic reticulum Ca2+ concentrations in a three-dimensional ER-PM junction and was solved using COMSOL Multiphysics. Modelling predicted that adhesion-dependent microdomains result from the concerted activity of IP3 receptors and pre-formed ORAI1-STIM2 complexes. In the present study, we extend this model to include the role of RYRs rapidly after TCR/CD3 stimulation. The involvement of STIM1, which has a lower KD for Ca2+ than STIM2, is also considered. Detailed 3D spatio-temporal simulations show that these Ca2+ microdomains rely on the concerted opening of ∼7 RYRs that are simultaneously active in response to the increase in NAADP induced by T cell stimulation. Opening of these RYRs provoke a local depletion of ER Ca2+ that triggers Ca2+ flux through the ORAI1 channels. Simulations predict that RYRs a, SCOPUS: ar.j, info:eu-repo/semantics/published

Embryonic stem cells (ESCs) are promising resources for both scientific research and clinical regenerative medicine. With regards to the latter, ESCs are especially useful for treating several neurodegenerative disorders. Two significant characteristics of ESCs, which make them so valuable, are their capacity for self-renewal and their pluripotency, both of which are regulated by the integration of various signaling pathways. Intracellular Ca2+ signaling is involved in several of these pathways. It is known to be precisely controlled by different Ca2+ channels and pumps, which play an important role in a variety of cellular activities, including proliferation, differentiation and apoptosis. Here, we provide a review of the recent work conducted to investigate the function of Ca2+ signaling in the self-renewal and the neural differentiation of ESCs. Specifically, we describe the role of intracellular Ca2+ mobilization mediated by RyRs (ryanodine receptors); by cADPR (cyclic adenosine 5'-diphosphate ribose) and CD38 (cluster of differentiation 38/cADPR hydrolase); and by NAADP (nicotinic acid adenine dinucleotide phosphate) and TPC2 (two pore channel 2). We also discuss the Ca2+ influx mediated by SOCs (store-operated Ca2+ channels), TRPCs (transient receptor potential cation channels) and LTCC (L-type Ca2+ channels) in the pluripotent ESCs as well as in neural differentiation of ESCs. Moreover, we describe the integration of Ca2+ signaling in the other signaling pathways that are known to regulate the fate of ESCs. (C) 2016 Elsevier Ltd. All rights reserved.

Embryonic stem cells (ESCs) are promising resources for both scientific research and clinical regenerative medicine. With regards to the latter, ESCs are especially useful for treating several neurodegenerative disorders. Two significant characteristics of ESCs, which make them so valuable, are their capacity for self-renewal and their pluripotency, both of which are regulated by the integration of various signaling pathways. Intracellular Ca2+ signaling is involved in several of these pathways. It is known to be precisely controlled by different Ca2+ channels and pumps, which play an important role in a variety of cellular activities, including proliferation, differentiation and apoptosis. Here, we provide a review of the recent work conducted to investigate the function of Ca2+ signaling in the self-renewal and the neural differentiation of ESCs. Specifically, we describe the role of intracellular Ca2+ mobilization mediated by RyRs (ryanodine receptors); by cADPR (cyclic adenosine 5'-diphosphate ribose) and CD38 (cluster of differentiation 38/cADPR hydrolase); and by NAADP (nicotinic acid adenine dinucleotide phosphate) and TPC2 (two pore channel 2). We also discuss the Ca2+ influx mediated by SOCs (store-operated Ca2+ channels), TRPCs (transient receptor potential cation channels) and LTCC (L-type Ca2+ channels) in the pluripotent ESCs as well as in neural differentiation of ESCs. Moreover, we describe the integration of Ca2+ signaling in the other signaling pathways that are known to regulate the fate of ESCs. (C) 2016 Elsevier Ltd. All rights reserved.

Embryonic stem cells (ESCs) are promising resources for both scientific research and clinical regenerative medicine. With regards to the latter, ESCs are especially useful for treating several neurodegenerative disorders. Two significant characteristics of ESCs, which make them so valuable, are their capacity for self-renewal and their pluripotency, both of which are regulated by the integration of various signaling pathways. Intracellular Ca2+ signaling is involved in several of these pathways. It is known to be precisely controlled by different Ca2+ channels and pumps, which play an important role in a variety of cellular activities, including proliferation, differentiation and apoptosis. Here, we provide a review of the recent work conducted to investigate the function of Ca2+ signaling in the self-renewal and the neural differentiation of ESCs. Specifically, we describe the role of intracellular Ca2+ mobilization mediated by RyRs (ryanodine receptors); by cADPR (cyclic adenosine 5'-diphosphate ribose) and CD38 (cluster of differentiation 38/cADPR hydrolase); and by NAADP (nicotinic acid adenine dinucleotide phosphate) and TPC2 (two pore channel 2). We also discuss the Ca2+ influx mediated by SOCs (store-operated Ca2+ channels), TRPCs (transient receptor potential cation channels) and LTCC (L-type Ca2+ channels) in the pluripotent ESCs as well as in neural differentiation of ESCs. Moreover, we describe the integration of Ca2+ signaling in the other signaling pathways that are known to regulate the fate of ESCs. (C) 2016 Elsevier Ltd. All rights reserved.

Various neurodegenerative disorders are associated with increased brain iron content. Iron is known to cause oxidative stress, which concomitantly promotes cell death. Whereas endolysosomes are known to serve as intracellular iron storage organelles, the consequences of increased iron on endolysosomal functioning, and effects on cell viability upon modulation of endolysosomal iron release remain largely unknown. Here, we show that increasing intracellular iron causes endolysosomal alterations associated with impaired autophagic clearance of intracellular protein aggregates, increased cytosolic oxidative stress and increased cell death. These effects are subject to regulation by NAADP, a potent second messenger reported to target endolysosomal TPCNs (2-pore channels). Consistent with endolysosomal iron storage, cytosolic iron levels are modulated by NAADP, and increased cytosolic iron is detected when overexpressing active, but not inactive TPCNs, indicating that these channels can modulate endolysosomal iron release. Cell death triggered by altered intralysosomal iron handling is abrogated in the presence of an NAADP antagonist or when inhibiting RAB7A activity. Taken together, our results suggest that increased endolysosomal iron causes cell death associated with increased cytosolic oxidative stress as well as autophagic impairments, and these effects are subject to modulation by endolysosomal ion channel activity in a RAB7A-dependent manner. These data highlight alternative therapeutic strategies for neurodegenerative disorders associated with increased intracellular iron load.

e release of Ca2+ from intracellular stores is a key process involved in a variety of signaling pathways. e second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) mobilizes Ca2+ from lysosomelike acidic stores but not from IP3-sensitive or ryanodine-sensitive Ca2+ stores in different cell types. To date, two receptors for NAADP have been discovered, namely TPC1 and TPC2. Interestingly, intracellular calcium dynamics plays a central role in angiogenesis of endothelial cells (EC) which is primarily triggered in response to growth factor such as vascular endothelial growth factor (VEGF). In addition to its well known role in the formation of new blood vessels, VEGF is also involved in a number of different EC functions such as proliferation, migration, regulation of vascular permeability and survival through the binding to two receptors (VEGFR1 and VEGFR2) both stimulating intracellular calcium mobilization. e aim of our study is to characterize the role of Ca2+ in VEGF-dependent signal transduction and to correlate its intracellular signaling to specific biological functions. Our data propose the involvement of NAADP in VEGFR2- but not VEGFR1-induced calcium mobilization. We show that both pharmacological inhibition of NAADP signaling with Ned-19 (a reported specific antagonist of NAADP) and the use of small interfering RNA (siRNA) targeting TPC1 and TPC2, significantly reduces VEGF-stimulated calcium release in Human Umbilical Vein Endothelial Cells (HUVEC). Our data suggest that both Ned-19 and silencing of TPC receptors by RNAi down regulates VEGF-stimulated ERK1/2 phosphorilation. Moreover, we observed that Ned-19 modulates Akt activation and inhibits VEGF-induced migration of EC. All together our data suggest that NAADP plays an important role in VEGF-induced Ca2+ signaling via tyrosine kinase receptor and open new strategies for studying EC functions.

A variety of endothelial agonist-induced responses are mediated by rises in intracellular Ca(2+), suggesting that different Ca(2+) signatures could fine-tune specific inflammatory and thrombotic activities. In search of new intracellular mechanisms modulating endothelial effector functions, we identified nicotinic acid adenine dinucleotide phosphate (NAADP) as a crucial second messenger in histamine-induced Ca(2+) release via H1 receptors (H1R). NAADP is a potent intracellular messenger mobilizing Ca(2+) from lysosome-like acidic compartments, functionally coupled to the endoplasmic reticulum. Using the human EA.hy926 endothelial cell line and primary human umbilical vein endothelial cells, we show that selective H1R activation increases intracellular NAADP levels and that H1R-induced calcium release involves both acidic organelles and the endoplasmic reticulum. To assess that NAADP links H1R to Ca(2+)-signaling we used both microinjection of self-inactivating concentrations of NAADP and the specific NAADP receptor antagonist, Ned-19, both of which completely abolished H1R-induced but not thrombin-induced Ca(2+) mobilization. Interestingly, H1R-mediated von Willebrand factor (VWF) secretion was completely inhibited by treatment with Ned-19 and by siRNA knockdown of 2-pore channel NAADP receptors, whereas thrombin-induced VWF secretion failed to be affected. These findings demonstrate a novel and specific Ca(2+)-signaling mechanism activated through H1R in human endothelial cells, which reveals an obligatory role of NAADP in the control of VWF secretion.

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a potent activator of intracellular Ca2+ release in several vertebrate and invertebrate systems. The role of the NAADP system in physiologicalprocesses is being extensively investigated at the present time. The NAADP receptor and its associated Ca2+ pool have been hypothesized to be important in several physiologicalprocesses including fertilization, T cell activation, and pancreaticsecretion. However, whether NAADP is a new second messenger ora tool for the discovery of a new Ca2+ channel is still an unansweredquestion. Research developed over the last two years has provided some important clues to whether NAADP is or not a physiological cellular messenger. In this short review, I will discuss some of these new findings that are helping us to find an answer to the important question: Is NAADP a second messenger or not?