Calcium Signaling In Response Research Articles

The synergistic effect of pulsed red light and leonurus inhibits primary dysmenorrhea induced by oxytocin in mice by modulating calcium signaling and inhibiting inflammatory responses.

Despite increasing evidence suggesting that red light photobiomodulation (R-PBM) and leonurus play important roles in analgesic and anti-inflammatory processes, data on their combined effect on primary dysmenorrhea (PD) are scarce. In this study, we reported the pain assessment of red light at various modes combined with leonurus on the oxytocin-induced model of PD mice. The combined intervention of pulsed R-PBM and leonurus decreased pain responses and PGF2α/PGE2 levels, alleviated uterine swelling and inflammatory infiltration, enhanced antioxidant levels (T-AOC, GSH-PX, SOD), and reduced lipid peroxidation (MDA, LPO) in the uterus, with its synergistic effect surpassing either treatment alone or the combination of continuous wave R-PBM with leonurus. Transcriptomic analysis demonstrated significant changes in differentially expressed genes associated with calcium signaling (Cav1, Cacna1c, Kcnmb1, Cnn1, and Myh11) and inflammatory response (Ptgs2, Jun, Fos, IL1rn, and IL17b) in the combination group, with concurrent downregulation of MLCK, COX-2, p-JNK/JNK, and IL17b protein levels, and upregulation of IL1rn, suggesting that the combined intervention of pulsed R-PBM and leonurus may alleviate pain through disruption of calcium homeostasis and induction of ROS-mediated inflammatory responses. Metabolomics studies of plasma revealed significant changes in lipid metabolism after the combined intervention, consistent with the transcriptomic findings. Hence, pulsed R-PBM combined leonurus has the potential to be an effective therapeutic approach for PD, as well as an alternative option for painful and inflammatory diseases; however, further exploration of its underlying mechanism is still necessary.

Robust In Vitro Models for Studying Parkinson's Disease? LUHMES Cells and SH-SH5Y Cells.

As our population ages, there is an increased unmet clinical need surrounding neurodegenerative diseases such as Parkinson's disease (PD). To tackle this ever-increasing problem, we must ensure that the cell models that we use to develop therapeutics in vitro are robust, reliable, and replicable. In this study, we compared SH-SY5Y cells with LUHMES cells in response to 6-Hydroxydopamine (6OHDA) and 1-Methyl-4-phenylpyridinium (MPP+), two common Parkinson's insults used in in vitro analysis. Both these cell types have apparent dopaminergic phenotypes, which could aid us in understanding their potential in this race to novel therapies. The LUHMES cells showed consistent dopaminergic (DA) expression through tyrosine hydroxylase (TH) positivity, alongside depleted ATP levels and elevated reactive oxygen species (ROS) production, whereas the SH-SH5Y cells displayed resilience to both chemical insults, raising questions about their utility in accurately modelling PD pathology. Further electrophysiological analysis revealed comparable firing rates and ion channel signalling between both cell types; however, LUHMES cells demonstrated stronger calcium signalling responses, further supporting their use as a more robust PD model. While SH-SY5Y cells showed some adaptability in vitro, their inconsistent DA phenotype and limited response to chemical insults limit their suitability for advanced research, suggesting that LUHMES cells should and must take their place as a hallmark in Parkinson's disease research.

Diatoms exhibit dynamic chloroplast calcium signals in response to high light and oxidative stress.

Diatoms are a group of silicified algae that play a major role in marine and freshwater ecosystems. Diatom chloroplasts were acquired by secondary endosymbiosis and exhibit important structural and functional differences from the primary plastids of land plants and green algae. Many functions of primary plastids, including photoacclimation and inorganic carbon acquisition, are regulated by calcium-dependent signaling processes. Calcium signaling has also been implicated in the photoprotective responses of diatoms; however, the nature of calcium elevations in diatom chloroplasts and their wider role in cell signaling remains unknown. Using genetically encoded calcium indicators, we find that the diatom Phaeodactylum tricornutum exhibits dynamic calcium elevations within the chloroplast stroma. Stromal calcium ([Ca2+]str) acts independently from the cytosol and is not elevated by stimuli that induce large cytosolic calcium ([Ca2+]cyt) elevations. In contrast, high light and exogenous hydrogen peroxide (H2O2) induce large, sustained [Ca2+]str elevations that are not replicated in the cytosol. Measurements using the fluorescent H2O2 sensor roGFP2-Oxidant Receptor Peroxidase 1 (Orp1) indicate that [Ca2+]str elevations induced by these stimuli correspond to the accumulation of H2O2 in the chloroplast. [Ca2+]str elevations were also induced by adding methyl viologen, which generates superoxide within the chloroplast, and by treatments that disrupt nonphotochemical quenching (NPQ). The findings indicate that diatoms generate specific [Ca2+]str elevations in response to high light and oxidative stress that likely modulate the activity of calcium-sensitive components in photoprotection and other regulatory pathways.

Beyond Wolfram Syndrome 1: The WFS1 Gene's Role in Alzheimer's Disease and Sleep Disorders.

The WFS1 gene was first identified in Wolfram Syndrome 1 (WS1), a rare autosomal recessive genetic disorder characterized by severe and progressive neurodegenerative changes. WFS1's role in various cellular mechanisms, particularly in calcium homeostasis and the modulation of endoplasmic reticulum (ER) stress, suggests its potential involvement in the pathogenesis of Alzheimer's disease (AD) and sleep disorders. Because it is involved in maintaining ER balance, calcium signaling, and stress responses, WFS1 plays a multifaceted role in neuronal health. Numerous studies have shown that the absence or improper expression of WFS1 disrupts these cellular processes, leading to neurodegeneration and making neurons more vulnerable. In AD, WFS1 dysfunction is thought to contribute to the accumulation of amyloid-β (Aβ) plaques and tau tangles, thereby accelerating disease progression. Additionally, WFS1 plays an essential role in sleep regulation by influencing neuronal excitability and neurotransmitter release, which may explain the sleep disturbances frequently observed in neurodegenerative diseases. Due to its involvement in the pathological mechanisms of AD and sleep disorders, WFS1 is regarded as a potential early diagnostic marker for these diseases. Further research is required to fully elucidate WFS1's role in the cellular pathway, perhaps facilitating the development of new therapeutic strategies for Alzheimer's disease and sleep disorders.

Osteocyte Sptbn1 Deficiency Alters Cell Survival and Mechanotransduction Following Formation of Plasma Membrane Disruptions (PMD) from Mechanical Loading.

We and others have shown that application of high-level mechanical loading promotes the formation of transient plasma membrane disruptions (PMD) which initiate mechanotransduction. We hypothesized that increasing osteocyte cell membrane fragility, by disrupting the cytoskeleton-associated protein β2-spectrin (Sptbn1), could alter osteocytic responses and bone adaptation to loading in a PMD-related fashion. In MLO-Y4 cells, treatment with the spectrin-disrupting agent diamide or knockdown of Sptbn1 via siRNA increased the number of PMD formed by fluid shear stress. Primary osteocytes from an osteocyte-targeted DMP1-Cre Sptbn1 conditional knockout (CKO) model mimicked trends seen with diamide and siRNA treatment and suggested the creation of larger PMD, which repaired more slowly, for a given level of stimulus. Post-wounding cell survival was impaired in all three models, and calcium signaling responses from the wounded osteocyte were mildly altered in Sptbn1 CKO cultures. Although Sptbn1 CKO mice did not demonstrate an altered skeletal phenotype as compared to WT littermates under baseline conditions, they showed a blunted increase in cortical thickness when subjected to an osteogenic tibial loading protocol as well as evidence of increased osteocyte death (increased lacunar vacancy) in the loaded limb after 2weeks of loading. The impaired post-wounding cell viability and impaired bone adaptation seen with Sptbn1 disruption support the existence of an important role for Sptbn1, and PMD formation, in osteocyte mechanotransduction and bone adaptation to mechanical loading.

Aequorin-Based In Vivo Luminescence Imaging Detects Calcium Signalling in Response to Biotic and Abiotic Stresses in Tomato

The tomato (Solanum lycopersicum L.), a widely cultivated and economically important vegetable crop, is subject to a number of biotic and abiotic stresses in nature. Several abiotic and biotic stresses have been demonstrated to elevate the concentration of cytosolic free Ca2+ ([Ca2+]i) in Arabidopsis due to the influx of calcium ions. In this study, recombinant aequorin was introduced into the tomato in order to investigate the change in [Ca2+]i when treated with exogenous Ca2+. This resulted in strong luminescence signals, which were mainly observed in the roots. Luminescence signals were also detected in the whole plant, including the leaves, when a surfactant (Silwet L-77) was added to coelenterazine. The concentration of [Ca2+]i increased with the dosage of NaCl/elf18. The luminescence signals also showed a lower increase in intensity with elf18 treatment compared to NaCl treatment. Furthermore, the [Ca2+]i responses to other abiotic or biotic stresses, such as H2O2 and Pep1, were also evaluated. It was found that this transgenic tomato expressing aequorin can effectively detect changes in [Ca2+]i levels. The transgenic tomato expressing aequorin represents an effective tool for detecting changes in [Ca2+]i and provides a solid basis for investigating the adaptation mechanisms of tomatoes to various abiotic and biotic stresses. Moreover, the aequorin-based system would be a highly valuable tool for studying the specificity and crosstalk of plant signalling networks under abiotic and biotic stresses in tomatoes.

Novel regulation of kidney function by gastrointestinal hormones

The control of whole-body energy metabolism involves intricate crosstalk between multiple organ systems including the gut and the brain that play well-recognized roles in this process. However, the important contributions and mechanisms by other organs are poorly understood. Errors in metabolic regulation can lead to human metabolic diseases including diabetes and obesity that affect 75% of the population and can lead to significant comorbidities. Improving our mechanistic understanding of multi-organ metabolic crosstalk in health and disease is essential for developing highly effcient and new therapeutic approaches. Macula densa (MD) cells in the kidney are chief regulators of renal blood flow and tissue remodeling. We hypothesized that the kidney is a major player in body metabolism via a new mechanism of gut-kidney crosstalk that involves the gut hormones gastrin and cholecystokinine (CCK) activating intrarenal, MD-specific mechanisms to increase blood flow and filtration, progenitor cells and tissue growth factors. High-resolution MD bulk and single-cell transcriptome analysis was performed from freshly isolated, enriched MD cells using MD-GFP mouse kidneys. Single cell MD Ca2+ signaling was analyzed in vivo using intravital multiphoton microscopy (MPM) of Sox2-GT mice that express the ratiometric Ca2+ reporter GCaMP6f/tdTomato in all renal cells. The altered expression of MD specific markers in response to gastrin/CCK8 stimuli was studied using western blot and the novel MDgeo immortalized cell line cultured in vitro. RNA sequencing and transcriptome analysis identified the gastrin/CCK receptor Cckbr as one of the highest expressed genes in MD versus control kidney cells. Intravital multiphoton imaging of Sox2-GCaMP6 reporter mouse kidneys revealed robust and entirely MD cell-specific calcium signaling in response to systemic gastrin/CCK injection. Using the novel MDgeo immortalized cell line cultured in vitro, CCK8 treatment led to 3-fold increases in MD cell synthesis of Prostaglandin E2 (PGE2)-generating COX2 and CCN1 that are established mediators of MD-induced glomerular hyperfiltration and tissue remodeling, respectively. Chronic treatment of Ren1d-Confetti mice with Darunavir, an HIV protease inhibitor significantly increased plasma CCK levels, Ren+ mesenchymal progenitor cell number and clonality, renal cell proliferation and tissue hypertrophy. This study uncovered new regulatory mechanisms of systemic metabolism especially the role of MD cells in the kidney. Repurposing the commonly used HIV drug Darunavir for kidney and metabolic diseases may provide therapeutic benefit for millions of patients worldwide. DK064234, DK123564, DK135290. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Increased Neuronal Nitric Oxide Synthase in Alzheimer's Disease Mediates Spontaneous Calcium Signaling and Divergent Glutamatergic Calcium Responses.

Nitrosative stress is a feature of Alzheimer's disease (AD). Aims: We aimed to identify the cause underpinning increased nitric oxide (NO) in neurons and the impact of NO on neuronal function in AD. Results: We analyzed neuronal nitric oxide synthase (nNOS) protein levels in postmortem tissue and induced pluripotent stem cell (iPSC)-derived neurons from Alzheimer's patients and controls by immunohistochemistry and Western blots. Furthermore, we assessed the impact of modulating nNOS function or NO levels on neuronal glutamatergic signaling using calcium imaging. We show that nNOS protein levels are increased in early and severely affected brain regions of AD postmortem tissue, but not late and mildly affected regions, or cognitively normal individuals. The increased nNOS phenotype was also present in iPSC-derived neurons from late-onset Alzheimer's disease (LOAD) patients compared with controls, along with increased levels of nitrite, a stable marker of NO. Innovation: We observed a divergent functional impact of NO that included strengthening the calcium response in control neurons, while dysregulating calcium signaling and altering the amplitude and kinetics of the calcium responses to glutamate in the AD neurons. Pharmacological scavenging of NO or inhibition of nNOS prevented aberrant spontaneous calcium signaling in AD neurons. Conclusion: Together these data identify increases in nNOS protein in AD. Functional data suggest that NO modulation of glutamatergic calcium signaling is neuroprotective under nonpathogenic conditions, with increased nNOS and NO contributing to dysregulated spontaneous calcium signaling in AD neurons.

Frequency-dependent characteristics of osteoblast calcium signaling responses to microvibrational stimulation

Frequency-dependent characteristics of osteoblast calcium signaling responses to microvibrational stimulation

VDR and PDIA3 Are Essential for Activation of Calcium Signaling and Membrane Response to 1,25(OH)2D3 in Squamous Cell Carcinoma Cells.

The genomic activity of 1,25(OH)2D3 is mediated by vitamin D receptor (VDR), whilst non-genomic is associated with protein disulfide isomerase family A member 3 (PDIA3). Interestingly, our recent studies documented that PDIA3 is also involved, directly or indirectly, in the modulation of genomic response to 1,25(OH)2D3. Moreover, PDIA3 was also shown to regulate cellular bioenergetics, possibly through the modulation of STAT signaling. Here, the role of VDR and PDIA3 proteins in membrane response to 1,25(OH)2D3 and calcium signaling was investigated in squamous cell carcinoma A431 cell line with or without the deletion of VDR and PDIA3 genes. Calcium influx was assayed by Fura-2AM or Fluo-4AM, while calcium-regulated element (NFAT) activation was measured using a dual luciferase assay. Further, the levels of proteins involved in membrane response to 1,25(OH)2D3 in A431 cell lines were analyzed via Western blot analysis. The deletion of either PDIA3 or VDR resulted in the decreased baseline levels of Ca2+ and its responsiveness to 1,25(OH)2D3; however, the effect was more pronounced in A431∆PDIA3. Furthermore, the knockout of either of these genes disrupted 1,25(OH)2D3-elicited membrane signaling. The data presented here indicated that the VDR is essential for the activation of calcium/calmodulin-dependent protein kinase II alpha (CAMK2A), while PDIA3 is required for 1,25(OH)2D3-induced calcium mobilization in A431 cells. Taken together, those results suggest that both VDR and PDIA3 are essential for non-genomic response to this powerful secosteroid.

Wild-type S100A3 and S100A13 restore calcium homeostasis and mitigate mitochondrial dysregulation in pulmonary fibrosis patient-derived cells

Patients with digenic S100A3 and S100A13 mutations exhibited an atypical and progressive interstitial pulmonary fibrosis, with impaired intracellular calcium homeostasis and mitochondrial dysfunction. Here we provide direct evidence of a causative effect of the mutation on receptor mediated calcium signaling and calcium store responses in control cells transfected with mutant S100A3 and mutant S100A13. We demonstrate that the mutations lead to increased mitochondrial mass and hyperpolarization, both of which were reversed by transfecting patient-derived cells with the wild type S100A3 and S100A13, or extracellular treatment with the recombinant proteins. In addition, we demonstrate increased secretion of inflammatory mediators in patient-derived cells and in control cells transfected with the mutant-encoding constructs. These findings indicate that treatment of patients’ cells with recombinant S100A3 and S100A13 proteins is sufficient to normalize most of cellular responses, and may therefore suggest the use of these recombinant proteins in the treatment of this devastating disease.

Difference in the Osteoblastic Calcium Signaling Response Between Compression and Stretching Mechanical Stimuli

In orthodontics, various forms of mechanical stimulation induce opposing bone metabolism mechanisms. Bone resorption and bone formation occur in areas of compressive and tensile force action, respectively. The mechanism that causes such a difference in bone metabolism is still unclear. In this study, we investigated the difference in the osteoblastic calcium signaling response between compression and stretching mechanical stimuli. We applied two types of mechanical stimuli to osteoblast-like MC3T3-E1 cells: first microneedle direct indentation onto the cell as compression stimuli, and second stretching stimuli by using originally developed cell stretching MEMS device. Cells were treated with thapsigargin and calcium-free medium to investigate the source of the calcium ion. The results demonstrated variations in the osteoblastic calcium signaling response between the compression and stretching stimuli. The magnitude of an increase in the intracellular calcium ion concentration is much higher in the compression stimuli-applied cell group. Treatment of calcium-free medium nearly suppressed the calcium signaling response to both types of mechanical stimulation. Thapsigargin treatment induced an increase in the magnitude of calcium signaling response to the compression stimuli, while suppressed the slow and sustained increase in the calcium ion concentration in the stretching stimuli-applied cell group. These findings demonstrate the difference in the characteristics of osteoblastic calcium signaling response between compression and stretching mechanical stimuli.

Ingredients of Vicks VapoRub inhibit rhinovirus-induced ATP release.

Over-the-counter therapies, such as Vicks VapoRub, are frequently used in the management of upper respiratory tract infection symptoms. Of these, acute cough is the most bothersome; however, the mechanisms involved have not been fully elucidated. The temperature-sensitive transient receptor potential (TRP) channels, including TRPA1, TRPV1, TRPM8 and TRPV4, are potential candidates. TRPV4 is also thought to be involved in cough through the TRPV4-ATP-P2X3 pathway. Here, we hypothesise that Vicks VapoRub ingredients (VVRIs) modulate the TRP cough channels. Stably transfected HEK cells expressing TRP channels were challenged with VVRIs, individually or in combination, and the agonist and antagonist effects were measured using calcium signalling responses. In addition, rhinovirus serotype-16 (RV16)-infected A549 airway epithelial cells were pre-incubated with individual or combinations of VVRIs prior to hypotonic challenge and extracellular ATP release analysis. Calcium signalling reconfirmed some previously defined activation of TRP channels by specific VVRIs. The combined VVRIs containing menthol, camphor and eucalyptus oil activated TRPV1, TRPV4, TRPM8 and untransfected wild-type HEK293 cells. However, pre-incubation with VVRIs did not significantly inhibit any of the channels compared with the standard agonist responses. Pre-incubation of RV16-infected A549 cells with individual or combined VVRIs, except menthol, resulted in a 0.45-0.55-fold reduction in total ATP release following hypotonic stimulation, compared with infected cells not treated with VVRIs. These findings suggest that some VVRIs may reduce symptoms associated with upper respiratory tract infection by modulating specific TRP receptors and by reducing RV16-induced ATP release.

Fiber optical imaging of astroglial calcium signaling in the respiratory network in the working heart brainstem preparation.

The neuronal activity in the respiratory network strongly depends on a variety of different neuromodulators. Given the essential role of astrocytes in stabilizing respiratory network activity generated by neurons in the preBötzinger complex (preBötC), our aim was to investigate astrocytic calcium signaling in the working heart brainstem preparation using fiber-optical imaging. By using transgenic mice that express GCaMP6s specifically in astrocytes, we successfully recorded astrocytic calcium signals in response to norepinephrine from individual astrocytes.

Role of Nitric Oxide in the Altered Calcium Homeostasis of Platelets from Rats with Biliary Cirrhosis.

Previously, we found that intracellular calcium (Ca2+) homeostasis is altered in platelets from an experimental model of liver cirrhosis, namely the bile-duct-ligated (BDL) rat. These alterations are compatible with the existence of a hypercoagulable state. In the present study, we analyzed the role of nitric oxide in the abnormal calcium signaling responses of an experimental cirrhosis model, the bile duct-ligated rat. Chronic treatment with L-NAME was used to inhibit NO production in a group of control and BDL animals, and the responses compared to those obtained in a control and BDL untreated group (n = 6 each). The experiments were conducted on isolated platelets loaded with fura-2, using fluorescence spectrometry. Chronic treatment with L-NAME increased thrombin-induced Ca2+ release from internal stores in both control and BDL rats. However, the effect was significantly greater in the BDL rats (p < 0.05). Thrombin-induced calcium entry from the extracellular space was also elevated but at lower doses and, similarly in both control and BDL platelets, treated with the NO synthesis inhibitor. Capacitative calcium entry was also enhanced in the control platelets but not in platelets from BDL rats treated with L-NAME. Total calcium in intracellular stores was elevated in untreated platelets from BDL rats, and L-NAME pretreatment significantly (p < 0.05) elevated these values both in controls and in BDL but significantly more in the BDL rats (p < 0.05). Our results suggest that nitric oxide plays a role in the abnormal calcium signaling responses observed in platelets from BDL rats by interfering with the mechanism that releases calcium from the internal stores.

A novel αB-crystallin R123W variant drives hypertrophic cardiomyopathy by promoting maladaptive calcium-dependent signal transduction.

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiovascular disorder affecting 1 in 500 people in the general population. Characterized by asymmetric left ventricular hypertrophy, cardiomyocyte disarray and cardiac fibrosis, HCM is a highly complex disease with heterogenous clinical presentation, onset and complication. While mutations in sarcomere genes can account for a substantial proportion of familial cases of HCM, 40%-50% of HCM patients do not carry such sarcomere variants and the causal mutations for their diseases remain elusive. Recently, we identified a novel variant of the alpha-crystallin B chain (CRYABR123W) in a pair of monozygotic twins who developed concordant HCM phenotypes that manifested over a nearly identical time course. Yet, how CRYABR123W promotes the HCM phenotype remains unclear. Here, we generated mice carrying the CryabR123W knock-in allele and demonstrated that hearts from these animals exhibit increased maximal elastance at young age but reduced diastolic function with aging. Upon transverse aortic constriction, mice carrying the CryabR123W allele developed pathogenic left ventricular hypertrophy with substantial cardiac fibrosis and progressively decreased ejection fraction. Crossing of mice with a Mybpc3 frame-shift model of HCM did not potentiate pathological hypertrophy in compound heterozygotes, indicating that the pathological mechanisms in the CryabR123W model are independent of the sarcomere. In contrast to another well-characterized CRYAB variant (R120G) which induced Desmin aggregation, no evidence of protein aggregation was observed in hearts expressing CRYABR123W despite its potent effect on driving cellular hypertrophy. Mechanistically, we uncovered an unexpected protein-protein interaction between CRYAB and calcineurin. Whereas CRYAB suppresses maladaptive calcium signaling in response to pressure-overload, the R123W mutation abolished this effect and instead drove pathologic NFAT activation. Thus, our data establish the CryabR123W allele as a novel genetic model of HCM and unveiled additional sarcomere-independent mechanisms of cardiac pathological hypertrophy.

Calcium-dependent activation of CPK12 facilitates its cytoplasm-to-nucleus translocation to potentiate plant hypoxia sensing by phosphorylating ERF-VII transcription factors

Calcium-dependent protein kinases (CDPKs/CPKs) are key regulators of plant stress signaling that translate calcium signals into cellular responses by phosphorylating diverse substrate proteins. However, the molecular mechanism by which plant cells relay calcium signals in response to hypoxia remains elusive. Here, we show that one member of the CDPK family in Arabidopsis thaliana, CPK12, is rapidly activated during hypoxia through calcium-dependent phosphorylation of its Ser-186 residue. Phosphorylated CPK12 shuttles from the cytoplasm to the nucleus, where it interacts with and phosphorylates the group VII ethylene-responsive transcription factors (ERF-VII) that are core regulators of plant hypoxia sensing, to enhance their stabilities. Consistently, CPK12 knockdown lines show attenuated tolerance of hypoxia, whereas transgenic plants overexpressing CPK12 display improved hypoxia tolerance. Nonethelss, loss of function of five ERF-VII proteins in an erf-vii pentuple mutant could partially suppress the enhanced hypoxia-tolerance phenotype of CPK12-overexpressing lines. Moreover, we also discovered that phosphatidic acid and 14-3-3κ protein serve as positive and negative modulators of the CPK12 cytoplasm-to-nucleus translocation, respectively. Taken together, these findings uncover a CPK12-ERF-VII regulatory module that is key to transducing calcium signals from the cytoplasm into the nucleus to potentiate hypoxia sensing in plants.

Local calcium signal transmission in mycelial network exhibits decentralized stress responses.

Many fungi live as mycelia, which are networks of hyphae. Mycelial networks are suited for the widespread distribution of nutrients and water. The logistical capabilities are critical for the extension of fungal survival areas, nutrient cycling in ecosystems, mycorrhizal symbioses, and virulence. In addition, signal transduction in mycelial networks is predicted to be vital for mycelial function and robustness. A lot of cell biological studies have elucidated protein and membrane trafficking and signal transduction in fungal hyphae; however, there are no reports visualizing signal transduction in mycelia. This paper, by using the fluorescent Ca2+ biosensor, visualized for the first time how calcium signaling is conducted inside the mycelial network in response to localized stimuli in the model fungus Aspergillus nidulans. The wavy propagation of the calcium signal inside the mycelium or the signal blinking in the hyphae varies depending on the type of stress and proximity to the stress. The signals, however, only extended around 1,500 μm, suggesting that the mycelium has a localized response. The mycelium showed growth delay only in the stressed areas. Local stress caused arrest and resumption of mycelial growth through reorganization of the actin cytoskeleton and membrane trafficking. To elucidate the downstream of calcium signaling, calmodulin, and calmodulin-dependent protein kinases, the principal intracellular Ca2+ receptors were immunoprecipitated and their downstream targets were identified by mass spectrometry analyses. Our data provide evidence that the mycelial network, which lacks a brain or nervous system, exhibits decentralized response through locally activated calcium signaling in response to local stress.

Loss of CFTR function is associated with reduced bitter taste receptor-stimulated nitric oxide innate immune responses in nasal epithelial cells and macrophages.

Bitter taste receptors (T2Rs) are G protein-coupled receptors identified on the tongue but expressed all over the body, including in airway cilia and macrophages, where T2Rs serve an immune role. T2R isoforms detect bitter metabolites (quinolones and acyl-homoserine lactones) secreted by gram negative bacteria, including Pseudomonas aeruginosa, a major pathogen in cystic fibrosis (CF). T2R activation by bitter bacterial products triggers calcium-dependent nitric oxide (NO) production. In airway cells, the NO increases mucociliary clearance and has direct antibacterial properties. In macrophages, the same pathway enhances phagocytosis. Because prior studies linked CF with reduced NO, we hypothesized that CF cells may have reduced T2R/NO responses, possibly contributing to reduced innate immunity in CF. Immunofluorescence, qPCR, and live cell imaging were used to measure T2R localization, calcium and NO signaling, ciliary beating, and antimicrobial responses in air-liquid interface cultures of primary human nasal epithelial cells and immortalized bronchial cell lines. Immunofluorescence and live cell imaging was used to measure T2R signaling and phagocytosis in primary human monocyte-derived macrophages. Primary nasal epithelial cells from both CF and non-CF patients exhibited similar T2R expression, localization, and calcium signals. However, CF cells exhibited reduced NO production also observed in immortalized CFBE41o- CF cells and non-CF 16HBE cells CRISPR modified with CF-causing mutations in the CF transmembrane conductance regulator (CFTR). NO was restored by VX-770/VX-809 corrector/potentiator pre-treatment, suggesting reduced NO in CF cells is due to loss of CFTR function. In nasal cells, reduced NO correlated with reduced ciliary and antibacterial responses. In primary human macrophages, inhibition of CFTR reduced NO production and phagocytosis during T2R stimulation. Together, these data suggest an intrinsic deficiency in T2R/NO signaling caused by loss of CFTR function that may contribute to intrinsic susceptibilities of CF patients to P. aeruginosa and other gram-negative bacteria that activate T2Rs.

Insight into calcium signalling in salt stress response

• SOS pathway is a major pathway triggered by Ca 2+ under salt stress. • Silicon has a role in the improved functioning of the SOS pathway under salt stress. • Calcium signalling and ABA synthesis results in stomatal closure. • High cytosolic Ca 2+ results in upregulating ROS production. Owing to the sessile nature of plants, they are at the mercy of environmental conditions and are often subject to severe abiotic stresses. Amongst these abiotic stresses, high salinity stress is one of the most common and severe stresses affecting one-third of irrigated agricultural lands worldwide. Calcium ion is well known for its essential role as a mediator of signals and in regulating and specifying the cellular responses to abiotic stresses. To adapt to salt stress, one of the main strategies developed by plants is calcium signalling responses. The excess extracellular salt is detected by specific root receptors and resulted in immediate rise of cytosolic free Ca 2+ . Exhaustive research has been conducted to identify the genetic identities of Ca 2+ channels and signalling factors, developing novel findings on such tolerance mechanisms in plants. However, the complex interconnected system of the salinity stress response by Ca 2+ signalling in plants has not been adequately justified. Plants employ a combination of Ca 2+ integrated proteins, phytohormones, osmolytes, receptors and other signalling factors in response to salinity stress, making this a complex calcium response network. In this review, we summarise and discuss the role of calcium signalling in response to salinity stress. In contrast to other studies, we try to provide a holistic overview of both chemical and mechanical processes involved in salt stress response while expanding on gaps filled by recent research. Furthermore, based on existing knowledge, we evaluate the different types of mechanisms and pathways through which calcium signalling works.