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1.
Cells ; 12(2)2023 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-36672231

RESUMO

Astrocytes' organisation affects the functioning and the fine morphology of the brain, both in physiological and pathological contexts. Although many aspects of their role have been characterised, their complex functions remain, to a certain extent, unclear with respect to their contribution to brain cell communication. Here, we studied the effects of nanotopography and microconfinement on primary hippocampal rat astrocytes. For this purpose, we fabricated nanostructured zirconia surfaces as homogenous substrates and as micrometric patterns, the latter produced by a combination of an additive nanofabrication and micropatterning technique. These engineered substrates reproduce both nanotopographical features and microscale geometries that astrocytes encounter in their natural environment, such as basement membrane topography, as well as blood vessels and axonal fibre topology. The impact of restrictive adhesion manifests in the modulation of several cellular properties of single cells (morphological and actin cytoskeletal changes) and the network organisation and functioning. Calcium wave signalling was observed only in astrocytes grown in confined geometries, with an activity enhancement in cells forming elongated agglomerates with dimensions typical of blood vessels or axon fibres. Our results suggest that calcium oscillation and wave propagation are closely related to astrocytic morphology and actin cytoskeleton organisation.


Assuntos
Astrócitos , Sinalização do Cálcio , Ratos , Animais , Astrócitos/metabolismo , Citoesqueleto/metabolismo , Microtúbulos/metabolismo , Hipocampo/metabolismo
2.
Science ; 379(6627): 66-71, 2023 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-36603091

RESUMO

Immotile cilia at the ventral node of mouse embryos are required for sensing leftward fluid flow that breaks left-right symmetry of the body. However, the flow-sensing mechanism has long remained elusive. In this work, we show that immotile cilia at the node undergo asymmetric deformation along the dorsoventral axis in response to the flow. Application of mechanical stimuli to immotile cilia by optical tweezers induced calcium ion transients and degradation of Dand5 messenger RNA (mRNA) in the targeted cells. The Pkd2 channel protein was preferentially localized to the dorsal side of immotile cilia, and calcium ion transients were preferentially induced by mechanical stimuli directed toward the ventral side. Our results uncover the biophysical mechanism by which immotile cilia at the node sense the direction of fluid flow.


Assuntos
Sinalização do Cálcio , Cálcio , Cílios , Mecanotransdução Celular , Animais , Camundongos , Cálcio/metabolismo , Cílios/fisiologia , Embrião de Mamíferos
3.
Science ; 379(6627): 71-78, 2023 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-36603098

RESUMO

The breaking of bilateral symmetry in most vertebrates is critically dependent upon the motile cilia of the embryonic left-right organizer (LRO), which generate a directional fluid flow; however, it remains unclear how this flow is sensed. Here, we demonstrated that immotile LRO cilia are mechanosensors for shear force using a methodological pipeline that combines optical tweezers, light sheet microscopy, and deep learning to permit in vivo analyses in zebrafish. Mechanical manipulation of immotile LRO cilia activated intraciliary calcium transients that required the cation channel Polycystin-2. Furthermore, mechanical force applied to LRO cilia was sufficient to rescue and reverse cardiac situs in zebrafish that lack motile cilia. Thus, LRO cilia are mechanosensitive cellular levers that convert biomechanical forces into calcium signals to instruct left-right asymmetry.


Assuntos
Padronização Corporal , Sinalização do Cálcio , Cálcio , Cílios , Peixe-Zebra , Animais , Cálcio/metabolismo , Cílios/fisiologia , Peixe-Zebra/crescimento & desenvolvimento , Proteínas de Peixe-Zebra/metabolismo , Canais de Cátion TRPP/metabolismo
4.
Cell Res ; 33(1): 71-79, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36588121

RESUMO

Pollen tube tip growth requires intricate Ca2+ signaling. Recent studies have also identified rapid alkalization factor (RALF)-family peptides and their receptors as critical components for pollen tube tip growth and integrity. The functional relationship of RALF and calcium signaling modules remains largely unclear. Here we report that disruption of RALF signaling pathway abolished the cytosolic Ca2+ gradient in the pollen tube, indicating that Ca2+ signaling is downstream of the RALF signaling pathway. We identified MILDEW RESISTANCE LOCUS O (MLO) family proteins MLO1, 5, 9, 15, as Ca2+ channels required for Ca2+ influx and pollen tube integrity. We further reconstituted the biochemical pathway in which signaling via RALF and RALF receptors activated MLO1/5/9/15 calcium channels. Together, we conclude that RALF peptides derived from pollen tube bind to their receptors to establish pollen tube Ca2+ gradient through activation of the MLO channels. Our finding has thus provided a mechanistic link between the RALF signaling pathway and Ca2+ signaling in controlling pollen tube integrity and growth.


Assuntos
Canais de Cálcio , Tubo Polínico , Tubo Polínico/metabolismo , Canais de Cálcio/metabolismo , Proteínas/metabolismo , Proteínas de Transporte/metabolismo , Peptídeos/metabolismo , Transdução de Sinais , Cálcio/metabolismo , Sinalização do Cálcio
5.
Biol Pharm Bull ; 46(1): 1-11, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36596517

RESUMO

Inspired by the well-known phenomenon of stretch-induced airway dilation in normal lungs and the emerging stretch-responsive Piezo1 channels that can be chemically activated by specific agonists such as Yoda1, we attempted to investigate whether chemical activation of Piezo1 by Yoda1 can modulate the biomechanical behaviors of airway smooth muscle cells (ASMCs) so that it may be exploited as a novel approach for bronchodilation. Thus, we treated in vitro cultured rat ASMCs with Yoda1, and examined the cells for calcium signaling, cell stiffness, traction force, cell migration, and the mRNA expression and distribution of molecules relevant to cell biomechanics. The data show that ASMCs expressed abundant mRNA of Piezo1. ASMCs exposed to 1 µM Yoda1 exhibited a potent but transient Ca2+ signaling, and treatment with 1 µM Yoda1 for 24 h led to decreased cell stiffness and traction force, all of which were partially reversed by Piezo1 inhibitor GsMTx4 and Piezo1 knockdown, respectively. In addition, ASMCs treated with 1 µM Yoda1 for 24 h exhibited impaired horizontal but enhanced vertical cell migration, as well as significant changes in key components of cells' contractile machinery including the structure and distribution of stress fibers and alpha-smooth muscle actin (α-SMA) fibrils, the mRNA expression of molecules associated with cell biomechanics. These results provide the first evidence that chemical activation of Piezo1 by Yoda1 resulted in marked pro-relaxation alterations of biomechanical behaviors and contractile machinery of the ASMCs. These findings suggest that Piezo1-specific agonists may indeed have great potential as alternative drug agents for relaxing ASMCs.


Assuntos
Sinalização do Cálcio , Miócitos de Músculo Liso , Ratos , Animais , Células Cultivadas , Miócitos de Músculo Liso/metabolismo , RNA Mensageiro/metabolismo
6.
Biochem Biophys Res Commun ; 643: 169-174, 2023 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-36610382

RESUMO

Dravet syndrome (DS) is an infantile-onset epileptic encephalopathy. More than 80% of DS patients have a heterozygous mutation in SCN1A, which encodes a subunit of the voltage-gated sodium channel, Nav1.1, in neurons. The roles played by astrocytes, the most abundant glial cell type in the brain, have been investigated in the pathogenesis of epilepsy; however, the specific involvement of astrocytes in DS has not been clarified. In this study, we evaluated Ca2+ signaling in astrocytes using genetically modified mice that have a loss-of-function mutation in Scn1a. We found that the slope of spontaneous Ca2+ spiking was increased without a change in amplitude in Scn1a+/- astrocytes. In addition, ATP-induced transient Ca2+ influx and the slope of Ca2+ spiking were also increased in Scn1a+/- astrocytes. These data indicate that perturbed Ca2+ dynamics in astrocytes may be involved in the pathogenesis of DS.


Assuntos
Epilepsias Mioclônicas , Epilepsia , Animais , Camundongos , Astrócitos/metabolismo , Epilepsias Mioclônicas/genética , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Canal de Sódio Disparado por Voltagem NAV1.1/metabolismo , Neurônios/metabolismo , Sinalização do Cálcio
7.
Int J Mol Sci ; 24(2)2023 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-36674826

RESUMO

In this study, we have investigated a possible mechanism that enables CB1/M3 receptor cross-talk, using SH-SY5Y cells as a model system. Our results show that M3 receptor activation initiates signaling that rapidly upregulates the CNR1 gene, resulting in a greatly potentiated CB1 receptor response to agonists. Calcium homeostasis plays an essential intermediary role in this functional CB1/M3 receptor cross-talk. We show that M3 receptor-triggered calcium release greatly increases CB1 receptor expression via both transcriptional and translational activity, by enhancing CNR1 promoter activity. The co-expression of M3 and CB1 receptors in brain areas such as the nucleus accumbens and amygdala support the hypothesis that the altered synaptic plasticity observed after exposure to cannabinoids involves cross-talk with the M3 receptor subtype. In this context, M3 receptors and their interaction with the cannabinoid system at the transcriptional level represent a potential pharmacogenomic target not only for the develop of new drugs for addressing addiction and tolerance. but also to understand the mechanisms underpinning response stratification to cannabinoids.


Assuntos
Canabinoides , Neuroblastoma , Humanos , Receptor CB1 de Canabinoide/genética , Receptor CB1 de Canabinoide/metabolismo , Cálcio/metabolismo , Canabinoides/farmacologia , Canabinoides/metabolismo , Sinalização do Cálcio
8.
Comput Biol Med ; 152: 106435, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36535207

RESUMO

BACKGROUND: Yuanjiang decoction (YJD), a traditional Chinese medicinal prescription, has been found to have a significant heart rate-increasing effect and is effective in the treatment of symptomatic bradyarrhythmia in previous studies. However, its specific components and potential mechanisms remain unclear. METHODS: In this study, we detected and identified the main compounds of YJD using liquid chromatography-mass spectrometry (LC-MS). Through the approach of network pharmacology, we predicted the core targets of the active components, bradyarrhythmia targets, and obtained potential anti-bradyarrhythmia targets of YJD. We further performed protein to protein interaction (PPI), gene ontology (GO) enrichment analyses and kyoto encyclopedia of genes and genomes (KEGG) signaling pathway analyses for core targets, and constructed network of key active ingredients-core targets of YJD. Finally, molecular docking and molecular dynamics simulation were performed for key active ingredients and core targets. RESULTS: The YJD contains a total of 35 main chemical components. The key active ingredients-core targets network contains 36 nodes and 90 edges, including 20 key active ingredients and 16 core targets. The core targets in the PPI network were TP53, TNF, HRAS, PPARG, IL1B, KCNH2, SCN5A, IDH1, LMNA, ACHE, F2, DRD2, CALM1, KCNQ1, TNNI3, IDH2 and TNNT2. KEGG pathway analysis showed that YJD treatment of bradyarrhythmia mainly involves neuroactive ligand-receptor interaction, adrenergic signaling in cardiomyocytes, cAMP signaling pathway, calcium signaling pathway, cholinergic synaptic and serotonergic synapse signaling pathway. The biological processes mainly include regulation of hormone levels, regulation of cardiac contraction, chemical synaptic transmission, circadian rhythm, positive regulation of heart rate, smooth muscle contraction, response to metal ion, oxidation-reduction process, neurotransmitter transport and import across plasma membrane. Molecular docking and molecular dynamics simulation results showed that hesperidin and tetrahydropalmatine had higher affinity with DRD2 and KCNQ1, respectively. CONCLUSION: This study reveals the pharmacodynamic material basis of YJD and its potential multicomponent-multitarget-multipathway pharmacological effects, predicted its potential anti-bradyarrhythmia mechanism may be related to the regulation of myocardial autonomic nervous function and related ion channels. Our work demonstrates that YJD has great potential for treating bradyarrhythmias as a complementary medicine, and the results can provide a theoretical basis for the development and clinical application of YJD.


Assuntos
Medicamentos de Ervas Chinesas , Farmacologia em Rede , Cromatografia Líquida , Canal de Potássio KCNQ1 , Simulação de Acoplamento Molecular , Espectrometria de Massas em Tandem , Sinalização do Cálcio , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Medicina Tradicional Chinesa
9.
Cell Immunol ; 383: 104651, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36493524

RESUMO

Lipopolysaccharides (LPS) is one of the most potent pathogen-associated signals for the immune system of vertebrates. In addition to the canonical pathway of LPS detection mediated by toll-like receptor 4 (TLR4) signaling pathway, TRP channel-mediated pathways endow sensory neurons and epithelial cells with the ability to detect and react to bacterial endotoxins. Previous work revealed that LPS triggers TRPV4-dependent calcium influx in urothelial cells (UCs) and mouse tracheobronchial epithelial cells (mTEC). In marked contrast, here we show that most subtypes of LPS could not directly activate TRPV4 channel. Although LPS from Salmonella enterica serotype Minnesota evoked a [Ca2+]i response in freshly isolated human bronchial epithelial cells (ECs), freshly isolated mouse ear skin single-cell suspensions, or HEK293T cells transiently transfected with mTRPV4, this activation occurred in a TRPV4-independent manner. Additionally, LPS from either E. coli strains or Salmonella enterica serotype Minnesota did not evoke significant difference in inflammation and pain hyperalgesia between wild type and TRPV4 deficient mice. In summary, our results demonstrate that in vitro and in vivo effects induced by LPS are independent of TRPV4, thus providing a clarity to the questioned role of LPS in TRPV4 activation.


Assuntos
Lipopolissacarídeos , Canais de Cátion TRPV , Humanos , Camundongos , Animais , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo , Canais de Cátion TRPV/farmacologia , Sinalização do Cálcio/fisiologia , Escherichia coli/metabolismo , Células HEK293 , Cálcio/metabolismo
10.
Neuropharmacology ; 222: 109306, 2023 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-36341808

RESUMO

Prolonged exposure (PE) therapy aiming to promote fear extinction is a useful treatment for post-traumatic stress disorder (PTSD). However, the mechanisms underlying fear extinction and effective methods used to promote fear extinction in PTSD are still lacking. In this study, we displayed dysfunctions of cyclic adenosine 3,5-monophosphate (cAMP)-protein kinase A (PKA), protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and calcium signaling in peripheral serum of PTSD patients using bioinformatics analysis. Later, we confirmed the dysfunctions of cAMP-PKA, AKT/mTOR and calcium signaling in the hippocampus of PTSD mice. Moreover, the reduction of calpain1 in the hippocampus enhanced fear memory acquisition. Single activation of PKA by systemic application of rolipram (ROL) or meglumine cyclic adenylate (M-cAMP) before re-exposure promoted fear extinction and improved anxiety-like behavior in PTSD mice. Moreover, systemic application of ROL before re-exposure improved hippocampal brain-derived neurotrophic factor (BDNF)/tyrosine kinase receptor B (TrkB) signaling and calpain1/AKT/mTOR signaling. Interestingly, the effects of activation of PKA could be partially blocked by TrkB antagonist, ANA-12 and mTOR inhibitor, RAPA. Finally, intranasal administration of ROL could also adjust the abnormality of fear memory and improve anxiety-like behaviors in PTSD mice. Collectively, activation of PKA could promote fear extinction, which correlated with the reduction of anxiety-like behavior. The mechanisms were related to the BDNF/TrkB and calpain1/AKT/mTOR signaling pathways. PKA activation might be a useful complementary therapy for PE in the symptom elimination of PTSD.


Assuntos
Medo , Transtornos de Estresse Pós-Traumáticos , Camundongos , Animais , Transtornos de Estresse Pós-Traumáticos/tratamento farmacológico , Proteínas Quinases Dependentes de AMP Cíclico , Proteínas Proto-Oncogênicas c-akt , Fator Neurotrófico Derivado do Encéfalo , Extinção Psicológica , Ansiedade/tratamento farmacológico , Serina-Treonina Quinases TOR , Rolipram , Sinalização do Cálcio , Adenosina , Mamíferos
11.
Nature ; 613(7942): 179-186, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36517594

RESUMO

Diffuse gliomas, particularly glioblastomas, are incurable brain tumours1. They are characterized by networks of interconnected brain tumour cells that communicate via Ca2+ transients2-6. However, the networks' architecture and communication strategy and how these influence tumour biology remain unknown. Here we describe how glioblastoma cell networks include a small, plastic population of highly active glioblastoma cells that display rhythmic Ca2+ oscillations and are particularly connected to others. Their autonomous periodic Ca2+ transients preceded Ca2+ transients of other network-connected cells, activating the frequency-dependent MAPK and NF-κB pathways. Mathematical network analysis revealed that glioblastoma network topology follows scale-free and small-world properties, with periodic tumour cells frequently located in network hubs. This network design enabled resistance against random damage but was vulnerable to losing its key hubs. Targeting of autonomous rhythmic activity by selective physical ablation of periodic tumour cells or by genetic or pharmacological interference with the potassium channel KCa3.1 (also known as IK1, SK4 or KCNN4) strongly compromised global network communication. This led to a marked reduction of tumour cell viability within the entire network, reduced tumour growth in mice and extended animal survival. The dependency of glioblastoma networks on periodic Ca2+ activity generates a vulnerability7 that can be exploited for the development of novel therapies, such as with KCa3.1-inhibiting drugs.


Assuntos
Neoplasias Encefálicas , Glioblastoma , Animais , Camundongos , Encéfalo/metabolismo , Encéfalo/patologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , NF-kappa B/metabolismo , Sistema de Sinalização das MAP Quinases , Sinalização do Cálcio , Morte Celular , Análise de Sobrevida , Cálcio/metabolismo
12.
Biochem Biophys Res Commun ; 642: 175-184, 2023 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-36584481

RESUMO

Dantrolene (DAN) directly binds to cardiac ryanodine receptor 2 (RyR2) through Leu601-Cys620 in the N-terminal domain and subsequently inhibits diastolic Ca2+ leakage through RyR2. We previously reported that therapy using RyR2 V3599K mutation, which inhibits diastolic Ca2+ leakage by enhancing calmodulin (CaM) binding ability to RyR2, prevents left ventricular (LV) remodeling in transverse aortic constriction (TAC) heart failure. Here, we examined whether chronic administration of DAN prevents LV remodeling in TAC heart failure via the same mechanism as genetic therapy. A pressure-overloaded hypertrophy mouse model was developed using TAC. Wild-type (WT) mice were divided into three groups: sham-operated mice (Sham group), TAC mice (TAC group), and TAC mice treated with DAN (TAC-DAN group, 20 mg/kg/day, i.p.). They were then followed up for 8 weeks. The survival rate was higher in the TAC-DAN group (83%) than in the TAC group (49%), and serial echocardiography studies and pathological tissue analysis showed that LV remodeling was significantly prevented in the TAC-DAN group compared to the TAC group. An increase in the diastolic Ca2+ spark frequency and a decrease in the binding affinity of CaM to RyR2 were observed at 8 weeks in the TAC group but not in the TAC-DAN group. Stabilization of RyR2 with DAN prevented LV remodeling and improved survival after TAC by enhancing CaM binding to RyR2 and inhibiting RyR2-mediated diastolic Ca2+ leakage.


Assuntos
Estenose da Valva Aórtica , Insuficiência Cardíaca , Camundongos , Animais , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Dantroleno/farmacologia , Dantroleno/uso terapêutico , Remodelação Ventricular/genética , Insuficiência Cardíaca/metabolismo , Sinalização do Cálcio
13.
FASEB J ; 37(1): e22683, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36520003

RESUMO

SIGIRR (single immunoglobulin IL-1 related receptor), PKP3 (plakophilin 3), and TMEM16J (anoctamin 9), a putative calcium-activated ion channel and phospholipid scramblase, control the immune response and the extent of inflammation. Variants of SIGIRR/PKP3/TMEM16J lead to severe inflammatory diseases such as pneumonia, enterocolitis, and kidney graft rejection. Meta-analysis of genome-wide association studies identified TMEM16J-T604A as a promotor for chronic kidney disease (CKD), but the disease mechanism and function of TMEM16J remain unknown. Here, we demonstrate TMEM16J as a calcium-activated calcium-permeable channel, which is expressed in the endoplasmic reticulum (ER). TMEM16J controls the intracellular distribution of calcium, and inhibits intracellular receptor-mediated Ca2+ signals and Ca2+ -dependent activation of ion channels, but augments transcription and release of pro-inflammatory cytokines. Renal epithelial cells expressing the variant TMEM16J-T604A show enhanced calcium signals when compared to cells expressing wt-TMEM16J, and demonstrate spontaneous transcription and release of cytokines. This study identifies TMEM16J as an important regulator of intracellular Ca2+ signals, ion channel activity, and cytokine release. TMEM16J may therefore affect immune response in renal tissue and immune cells.


Assuntos
Cálcio , Estudo de Associação Genômica Ampla , Cálcio/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Canais de Cálcio/metabolismo , Receptores de Interleucina-1/genética , Citocinas , Sinalização do Cálcio/fisiologia
14.
Pancreatology ; 23(1): 1-8, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36539315

RESUMO

In this account of the 2022 Palade Medal Lecture, an attempt is made to explain, as simply as possible, the most essential features of normal physiological control of pancreatic enzyme secretion, as they have emerged from more than 50 years of experimental work. On that basis, further studies on the mechanism by which acute pancreatitis is initiated are then described. Calcium ion signaling is crucially important for both the normal physiology of secretion control as well as for the development of acute pancreatitis. Although acinar cell processes have, rightly, been central to our understanding of pancreatic physiology and pathophysiology, attention is here drawn to the additional critical influence of calcium signaling events in stellate and immune cells in the acinar environment. These signals contribute significantly to the crucially important inflammatory response in acute pancreatitis.


Assuntos
Distinções e Prêmios , Pancreatite , Humanos , Doença Aguda , Sinalização do Cálcio , Células Acinares/metabolismo , Cálcio/metabolismo
15.
Glia ; 71(1): 44-59, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-35822691

RESUMO

The study of the astrocytic contribution to brain functions has been growing in popularity in the neuroscience field. In the last years, and especially since the demonstration of the involvement of astrocytes in synaptic functions, the astrocyte field has revealed multiple functions of these cells that seemed inconceivable not long ago. In parallel, cannabinoid investigation has also identified different ways by which cannabinoids are able to interact with these cells, modify their functions, alter their communication with neurons and impact behavior. In this review, we will describe the expression of different endocannabinoid system members in astrocytes. Moreover, we will relate the latest findings regarding cannabinoid modulation of some of the most relevant astroglial functions, namely calcium (Ca2+ ) dynamics, gliotransmission, metabolism, and inflammation.


Assuntos
Astrócitos , Canabinoides , Astrócitos/metabolismo , Endocanabinoides/metabolismo , Neurônios/metabolismo , Cálcio/metabolismo , Sinalização do Cálcio/fisiologia
16.
Cell Calcium ; 109: 102675, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36525777

RESUMO

NAADP (nicotinic acid adenine dinucleotide phosphate) is a second messenger, releasing Ca2+ from acidic calcium stores such as endosomes and lysosomes. PI(3,5)P2 (phosphatidylinositol 3,5-bisphosphate) is a phospho-inositide, residing on endolysosomal membranes and likewise releasing Ca2+ from endosomes and lysosomes. Both compounds have been shown to activate endolysosomal two-pore channels (TPCs) in mammalian cells. However, their effects on ion permeability as demonstrated specifically for TPC2 differ. While PI(3,5)P2 elicits predominantly Na+-selective currents, NAADP increases the Ca2+ permeability of the channel. What happens when both compounds are applied simultaneously was unclear until recently.


Assuntos
Canais de Cálcio , Sinalização do Cálcio , Animais , Canais de Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , NADP/metabolismo , Cálcio/metabolismo , Lisossomos/metabolismo , Mamíferos/metabolismo
17.
Cell Calcium ; 109: 102688, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36538845

RESUMO

Contact sites between the endoplasmic reticulum (ER) and mitochondria play a pivotal role in cell signaling, and the interaction between these organelles is dynamic and finely regulated. We have studied the role of ER Ca2+ concentration ([Ca2+]ER) in modulating this association in HeLa and HEK293 cells and human fibroblasts. According to Manders' coefficient, ER-mitochondria colocalization varied depending on the ER marker; it was the highest with ER-Tracker and the lowest with ER Ca2+ indicators (Mag-Fluo-4, erGAP3, and G-CEPIA1er) in both HeLa cells and human fibroblasts. Only GEM-CEPIA1er displayed a high colocalization with elongated mitochondria in HeLa cells, this ER Ca2+ indicator reveals low Ca2+ regions because this ion quenches its fluorescence. On the contrary, the typical rounded and fragmented mitochondria of HEK293 cells colocalized with Mag-Fluo-4 and, to a lesser extent, with GEM-CEPIA1er. The ablation of the three IP3R isoforms in HEK293 cells increased mitochondria-GEM-CEPIA1er colocalization. This pattern of colocalization was inversely correlated with the rate of ER Ca2+ leak evoked by thapsigargin (Tg). Moreover, Tg and Histamine in the absence of external Ca2+ increased mitochondria-ER colocalization. On the contrary, in the presence of external Ca2+, both Bafilomycin A1 and Tg reduced the mitochondria-ER interaction. Notably, knocking down MCU decreased mitochondria-ER colocalization. Overall, our data suggest that the [Ca2+] is not homogenous within the ER lumen and that mitochondria-ER interaction is modulated by the ER Ca2+ leak and the [Ca2+]i.


Assuntos
Retículo Endoplasmático , Mitocôndrias , Humanos , Células HeLa , Células HEK293 , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Tapsigargina/farmacologia , Cálcio/metabolismo , Sinalização do Cálcio
18.
Trends Plant Sci ; 28(1): 74-89, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36504136

RESUMO

Calcium ions (Ca2+) are prominent intracellular messengers in all eukaryotic cells. Recent studies have emphasized the crucial roles of Ca2+ in plant immunity. Here, we review the latest progress on the spatiotemporal control of Ca2+ function in plant immunity. We discuss discoveries of how Ca2+ influx is triggered upon the activation of immune receptors, how Ca2+-permeable channels are activated, how Ca2+ signals are decoded inside plant cells, and how these signals are switched off. Despite recent advances, many open questions remain and we highlight the existing toolkit and the new technologies to address the outstanding questions of Ca2+ signaling in plant immunity.


Assuntos
Sinalização do Cálcio , Cálcio , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Imunidade Vegetal/genética , Células Vegetais/metabolismo
19.
J Agric Food Chem ; 70(51): 16156-16163, 2022 Dec 28.
Artigo em Inglês | MEDLINE | ID: mdl-36524829

RESUMO

Ryanodine receptor (RyR) is a giant calcium release channel located on the membrane of the endoplasmic reticulum (ER). Here, we report the regulation of RyRs from two major agricultural pests, diamondback moth and fall armyworm, by insect calmodulin (CaM). The recombinantly expressed full-length insect RyR could be pulled down by insect CaM in the presence of Ca2+, but the efficiency is lower compared to rabbit RyR1 and insect RyR with the CaM-binding domain (CaMBD) replaced by rabbit RyR1 sequence. Interestingly, the enhanced binding of CaM in the mutant insect RyR resulted in an increased sensitivity to the diamide insecticide chlorantraniliprole (CHL), suggesting that this CaM-CaMBD interface could be targeted by potential synergists acting as molecular glue. The thermodynamics of the binding between insect CaM and CaMBD was characterized by isothermal titration calorimetry, and the key residues responsible for the insect-specific regulation were identified through mutagenesis studies.


Assuntos
Calmodulina , Mariposas , Animais , Coelhos , Calmodulina/genética , Calmodulina/química , Calmodulina/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Mariposas/genética , Mariposas/metabolismo , Sinalização do Cálcio , Ligação Proteica , Cálcio/metabolismo
20.
Cell Death Dis ; 13(12): 1063, 2022 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-36543780

RESUMO

Epilepsy is a common neurological disorder and glutamate excitotoxicity plays a key role in epileptic pathogenesis. Astrocytic glutamate transporter GLT-1 is responsible for preventing excitotoxicity via clearing extracellular accumulated glutamate. Previously, three variants (G82R, L85P, and P289R) in SLC1A2 (encoding GLT-1) have been clinically reported to be associated with epilepsy. However, the functional validation and underlying mechanism of these GLT-1 variants in epilepsy remain undetermined. In this study, we reported that these disease-linked mutants significantly decrease glutamate uptake, cell membrane expression of the glutamate transporter, and glutamate-elicited current. Additionally, we found that these variants may disturbed stromal-interacting molecule 1 (STIM1)/Orai1-mediated store-operated Ca2+ entry (SOCE) machinery in the endoplasmic reticulum (ER), in which GLT-1 may be a new partner of SOCE. Furthermore, knock-in mice with disease-associated variants showed a hyperactive phenotype accompanied by reduced glutamate transporter expression. Therefore, GLT-1 is a promising and reliable therapeutic target for epilepsy interventions.


Assuntos
Cálcio , Epilepsia , Camundongos , Animais , Molécula 1 de Interação Estromal , Cálcio/metabolismo , Epilepsia/genética , Ácido Glutâmico/metabolismo , Transporte Biológico , Proteína ORAI1/metabolismo , Sinalização do Cálcio
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