Calcium-permeable Channel Research Articles

Evolutionary analysis of the OSCA gene family in sunflower (Helianthus annuus L) and expression analysis under NaCl stress.

Hyperosmolality-gated calcium-permeable channels (OSCA) are Ca2+ nonselective cation channels that contain the calcium-dependent DUF221 domain, which plays an important role in plant response to stress and growth. However, the OSCA gene has not been fully identified and analyzed in sunflowers. In this study, we comprehensively analyzed the number, structure, collinearity, and phylogeny of the OSCA gene family in the sunflower, six Compositae species (Arctium lappa, Chrysanthemum morifolium, Cichorium endivia, Cichorium intybus, Lactuca sativa var. Angustata, and Carthamus tinctorius), and six other plants (soybean, Arabidopsis thaliana, rice, grape, and maize). The expression of the sunflower OSCA gene in nine different tissues, six different hormones, and NaCl stress conditions were analyzed based on transcriptome data and qRT-PCR. A total of 15 OSCA proteins, distributed on 10 chromosomes, were identified in the sunflower, and all of them were located in the endoplasmic reticulum. Using the phylogenetic tree, collinearity, gene structure, and motif analysis of the six Compositae species and six other plants, we found that the sunflower OSCA protein had only three subfamilies and lacked the Group 4 subfamily, which is conserved in the evolution of Compositae and subject to purification selection. The OSCA gene structure and motif analysis of the sunflower and six Compositae showed that there was a positive correlation between the number of motifs of most genes and the length of the gene, different subfamilies had different motifs, and the Group 4 subfamily had the smallest number of genes and the simplest gene structure. RNA-seq and qRT-PCR analysis showed that the expression levels of most OSCA genes in the sunflower changed to varying degrees under salt stress, and HaOSCA2.6 and HaOSCA3.1 were the most important in the sunflower's response to salt stress. The coexpression network of the sunflower genes under salt stress was constructed based on weighted gene co-expression network analysis (WGCNA). In conclusion, our findings suggest that the OSCA gene family is conserved during the sunflower's evolution and plays an important role in salt tolerance. These results will deepen our understanding of the evolutionary relationship of the sunflower OSCA gene family and provide a basis for their functional studies under salt stress.

Genome-wide characterization and comparative analysis of the OSCA gene family and identification of its potential stress-responsive members in legumes

Cicer arietinum, Cajanus cajan, Vigna radiata, and Phaseolus vulgaris are economically important legume crops with high nutritional value. They are negatively impacted globally by different biotic and abiotic stresses. Hyperosmolality-gated calcium-permeable channels (OSCA) have been characterized as osmosensors in Arabidopsis thaliana but have not previously reported in legumes. This study provides a genome-wide identification, characterization, and comparative analysis of OSCA genes in legumes. Our study identified and characterized 13 OSCA genes in C. cajan, V. radiata, P. vulgaris, and 12 in C. arietinum, classified into four distinct clades. We found evidence to suggest that the OSCAs might be involved in the interaction between hormone signalling pathways and stress signalling pathways. Furthermore, they play a major role in plant growth and development. The expression levels of the OSCAs vary under different stress conditions in a tissue-specific manner. Our study can be used to develop a detailed understanding of stress regulatory mechanisms of the OSCA gene family in legumes.

IPLA2 inhibition blocks LysoPC-induced TRPC6 externalization and promotes Re-endothelialization of carotid injuries in hypercholesterolemic mice

Lipid oxidation products, including lysophosphatidylcholine (lysoPC), accumulate at the site of arterial injury after vascular interventions and hinder re-endothelization. LysoPC activates calcium-permeable channels, specifically canonical transient receptor potential 6 (TRPC6) channels that induce a sustained increase in intracellular calcium ion concentration [Ca2+]i and contribute to dysregulation of the endothelial cell (EC) cytoskeleton. Activation of TRPC6 leads to inhibition of EC migration in vitro and delayed re-endothelization of arterial injuries in vivo. Previously, we demonstrated the role of phospholipase A2 (PLA2), specifically calcium-independent PLA2 (iPLA2), in lysoPC-induced TRPC6 externalization and inhibition of EC migration in vitro. The ability of FKGK11, an iPLA2-specific pharmacological inhibitor, to block TRPC6 externalization and preserve EC migration was assessed in vitro and in a mouse model of carotid injury. Our data suggest that FKGK11 prevents lysoPC-induced PLA2 activity, blocks TRPC6 externalization, attenuates calcium influx, and partially preserves EC migration in vitro. Furthermore, FKGK11 promotes re-endothelization of an electrocautery carotid injury in hypercholesterolemic mice. FKGK11 has similar arterial healing effects in male and female mice on a high-fat diet. This study suggests that iPLA2 is a potential therapeutic target to attenuate calcium influx through TRPC6 channels and promote EC healing in cardiovascular patients undergoing angioplasty.

Suppression of highly efficient cadmium translocation by exogenous gibberellin provides insights into the molecular regulation mechanism of hyperaccumulation in Sedum plumbizincicola

Sedum plumbizincicola is a hyperaccumulating species with highly efficient uptake, translocation and distribution system for cadmium (Cd), a non-essential and toxic heavy metal. To understand the molecular regulation mechanisms of Cd transport and accumulation in this hyperaccumulator is still a challenge. In this study, we observed that exogenous gibberellin A3 (GA3) significantly reduced Cd loading efficiency from root to shoot and uploading efficiency from stem to leaves in S. plumbizincicola under Cd treatments, which provided us a remarkable opportunity to study the signaling mechanisms for regulation of highly efficient Cd transport and translocation in S. plumbizincicola. Here, we constructed a de novo full-length transcriptome database with the integration of full-length and next-generation transcriptome sequences for Cd-treated S. plumbizincicola with or without exogenous GA3 application. From this database, we identified 1461 differentially expressed genes (DEGs), which were related to solute transport, cell wall metabolism and signal transduction, and most of them were down-regulated by the exogenous application of gibberellin. In particularly, a substantial number of the downregulated genes belong to calmodulin-like (CML) proteins, calcium-dependent protein kinase, calcium-permeable channel, mitochondrial calcium uniporter (MCU) and Na+/Ca2+ exchanger-like proteins (NCL/EF-CAX). These results indicated that calcium homeostasis and signal transduction may play an important role in the regulation of highly efficient translocation of cadmium in hyperaccumulator S. pumbizincicola.

Negative self-regulation of transient receptor potential canonical 4 by the specific interaction with phospholipase C-δ1

Transient receptor potential canonical (TRPC) channels are non-selective calcium-permeable cation channels. It is suggested that TRPC4β is regulated by phospholipase C (PLC) signaling and is especially maintained by phosphatidylinositol 4,5-bisphosphate (PIP2). In this study, we present the regulation mechanism of the TRPC4 channel with PIP2 hydrolysis which is mediated by a channel-bound PLCδ1 but not by the GqPCR signaling pathway. Our electrophysiological recordings demonstrate that the Ca2+ via an open TRPC4 channel activates PLCδ1 in the physiological range, and it causes the decrease of current amplitude. The existence of PLCδ1 accelerated PIP2 depletion when the channel was activated by an agonist. Interestingly, PLCδ1 mutants which have lost the ability to regulate PIP2 level failed to reduce the TRPC4 current amplitude. Our results demonstrate that TRPC4 self-regulates its activity by allowing Ca2+ ions into the cell and promoting the PIP2 hydrolyzing activity of PLCδ1.

Transient receptor potential melastatin 2 regulates neutrophil extracellular traps formation and delays resolution of neutrophil-driven sterile inflammation

The formation of neutrophil extracellular traps (NETs) is a process releasing into the extracellular space networks of chromatin fibers decorated with granular proteins. It is implicated in infection-related as well as sterile inflammation. Monosodium urate (MSU) crystals serve as damage-associated molecular pattern (DAMP) in various conditions of disease. Formation of NETs or aggregated NETs (aggNETs) orchestrates initiation and resolution of MSU crystals-triggered inflammation, respectively. Elevated intracellular calcium levels and the generation of reactive oxygen species (ROS) are crucial for the formation of MSU crystal-induced NETs. However, the exact signaling pathways involved are still elusive. Herein, we demonstrate that the ROS-sensing, non-selective calcium-permeable channel transient receptor potential cation channel subfamily M member 2 (TRPM2) is required for a full-blown MSU crystal-induced NET formation. Primary neutrophils from TRPM2−/− mice showed reduced calcium influx and ROS production and, consequently a reduced formation of MSU crystal-induced NETs and aggNETs. Furthermore, in TRPM2−/− mice the infiltration of inflammatory cells into infected tissues and their production of inflammatory mediators was suppressed. Taken together these results describe an inflammatory role of TRPM2 for neutrophil-driven inflammation and identify TRPM2 as potential target for therapeutic intervention.

Spinophilin-dependent regulation of GluN2B-containing NMDAR-dependent calcium influx, GluN2B surface expression, and cleaved caspase expression.

N-methyl-d-aspartate receptors (NMDARs) are calcium-permeable ion channels that are ubiquitously expressed within the glutamatergic postsynaptic density. Phosphorylation of NMDAR subunits defines receptor conductance and surface localization, two alterations that can modulate overall channel activity. Modulation of NMDAR phosphorylation by kinases and phosphatases regulates the amount of calcium entering the cell and subsequent activation of calcium-dependent processes. The dendritic spine enriched protein, spinophilin, is the major synaptic protein phosphatase 1 (PP1) targeting protein. Depending on the substrate, spinophilin can act as either a PP1 targeting protein, to permit substrate dephosphorylation, or a PP1 inhibitory protein, to enhance substrate phosphorylation. Spinophilin limits NMDAR function in a PP1-dependent manner. Specifically, we have previously shown that spinophilin sequesters PP1 away from the GluN2B subunit of the NMDAR, which results in increased phosphorylation of Ser-1284 on GluN2B. However, how spinophilin modifies NMDAR function is unclear. Herein, we utilize a Neuro2A cell line to detail that Ser-1284 phosphorylation increases calcium influx via GluN2B-containing NMDARs. Moreover, overexpression of spinophilin decreases GluN2B-containing NMDAR activity by decreasing its surface expression, an effect that is independent of Ser-1284 phosphorylation. In hippocampal neurons isolated from spinophilin knockout animals, there is an increase in cleaved caspase-3 levels, a marker of calcium-associated apoptosis, compared with wildtype mice. Taken together, our data demonstrate that spinophilin regulates GluN2B containing NMDAR phosphorylation, channel function, and trafficking and that loss of spinophilin enhances neuronal cleaved caspase-3 expression.

Gsmtx4 Alleviated Osteoarthritis through Piezo1/Calcineurin/NFAT1 Signaling Axis under Excessive Mechanical Strain.

Excessive mechanical strain is the prominent risk factor for osteoarthritis (OA), causing cartilage destruction and degeneration. However, the underlying molecular mechanism contributing to mechanical signaling transduction remains unclear in OA. Piezo type mechanosensitive ion channel component 1 (Piezo1) is a calcium-permeable mechanosensitive ion channel and provides mechanosensitivity to cells, but its role in OA development has not been determined. Herein, we found up-regulated expression of Piezo1 in OA cartilage, and that its activation contributes to chondrocyte apoptosis. The knockdown of Piezo1 could protect chondrocytes from apoptosis and maintain the catabolic and anabolic balance under mechanical strain. In vivo, Gsmtx4, a Piezo1 inhibitor, markedly ameliorated the progression of OA, inhibited the chondrocyte apoptosis, and accelerated the production of the cartilage matrix. Mechanistically, we observed the elevated activity of calcineurin (CaN) and the nuclear transfection of nuclear factor of activated T cells 1 (NFAT1) under mechanical strain in chondrocytes. Inhibitors of CaN or NFAT1 rescued the pathologic changes induced by mechanical strain in chondrocytes. Overall, our findings revealed that Piezo1 was the essential molecule response to mechanical signals and regulated apoptosis and cartilage matrix metabolism via the CaN/NFAT1 signaling axis in chondrocytes, and that Gsmtx4 could be an attractive therapeutic drug for OA treatment.

Calcium-dependent protein kinase CDPK16 phosphorylates serine-856 of glutamate receptor-like GLR3.6 protein leading to salt-responsive root growth in Arabidopsis.

Calcium-permeable channels in the plasma membrane play vital roles in plant growth, development, and response to environmental stimuli. Arabidopsis possesses 20 glutamate receptor-like proteins that share similarities with animal ionotropic glutamate receptors and mediate Ca2+ influx in plants. Calcium-dependent protein kinases (CDPKs) phosphorylate serine (Ser)-860 of glutamate receptor-like (GLR)3.7 protein, which interacts with 14-3-3ω and plays an essential role in salt and abscisic acid response in Arabidopsis by modulating Ca2+ signaling. However, the significance of CDPK- mediated phosphorylation status of Ser residues of GLR3.6 with regard to the functioning of GLR3.6 remains to be elucidated. In this study, we performed an in vitro kinase assay using CDPK16 and peptides containing the 14-3-3ω interacting domain of GLR3.6. We showed that Ser861/862 of GLR3.6 are required for the interaction with 14-3-3ω and that Ser856 of GLR3.6 is specifically phosphorylated by CDPK16 but not by CDPK3 and CDPK34. In addition, the expression of GLR3.6 was quickly downregulated by salt stress, and plants of glr3.6 mutants and GLR3.6-overexpression lines presented shorter and longer root lengths, respectively, under normal growth conditions than Col. Overexpression of the GLR3.6-Ser856 to Ala mutation resulted in a less sensitive phenotype in response to salt stress similar to glr3.6. Our results indicated that the Ser861/862 residues of GLR3.6 are required for interaction with 14-3-3ω. Additionally, the phosphorylation status of Ser856 residue of GLR3.6, which is mediated specifically by CDPK16, regulates root growth in normal and salt stress and conditions.

Abstract WP232: Endothelial Knockout Of Piezo1 Results In Reduced Cerebral Blood Flow, Brain Gliosis, And Functional Changes In Gait.

Introduction: Piezo1 is a calcium-permeable mechanosensitive ion channel that is opened by membrane stretch or increased shear force. In peripheral vasculature, activation of endothelial Piezo1 promotes vasodilation. Whether endothelial Piezo1 regulates cerebral blood flow and the consequences of endothelial cell (EC) Piezo1 loss-of-function in the brain are unknown. The present study was designed to test the overall hypothesis that loss of EC Piezo1 results in reduced cerebral blood flow and subsequent brain pathology consistent with chronic hypoperfusion. Methods: EC specific Piezo1 knockout mice were generated by crossing Piezo1 fl/fl mice with inducible EC-specific Cre mice (Cdh5(Pac)CreERT2). Tamoxifen (75 mg/kg) was administrated to generate EC Piezo1 KO or as control in Piezo1 “floxed only” mice. At 1 month after tamoxifen injection, three cohorts were evaluated: (cohort 1) measure CBF by MRI ASL, (cohort 2) gait analysis (Digigait) and subsequent immunofluorescence (IF), (cohort 3) scRNAseq analysis. Gait analysis was chosen as a behavioral measure sensitive to white matter injury. Results: EC Piezo1 KO resulted in >8% reduction in resting CBF in striatum (n=5; p<0.05, females). Gait analysis showed significant change in stride time, stride length, and cadence (n = 9-10 males, 6-9 females; p< 0.01). IF showed gliosis in the corpus callosum of EC Piezo1 KO mice, in the form of astrogliosis (Gfap) in males and microgliosis (Iba1) in females (n=5-7 in each group; p<0.05). From scRNAseq of EC Piezo1 KO and control brains (males), GO analysis from all clusters revealed GO terms including: inflammatory response, synapse pruning, gliogenesis, learning or memory, and locomotory behavior. GO analysis from EC clusters revealed GO terms including: gliogenesis, myelination, and vascular morphogenesis. Among the most highly upregulated genes in the EC cluster, were Ctss and C1q family genes. In choroid plexus cluster, EC Piezo1 KO showed increased expression of Hif1a target genes. Conclusions: We conclude that EC Piezo1 loss-of-function promotes hypoperfusion sufficient to induce induction of inflammatory signaling and hypoxia-dependent transcription. White matter injury was evident as gliosis in the corpus callosum and by alterations in gait.

TRP434 and TRP435 residues are crucial for sensing calcium ions and PI(4,5)P2 molecules in TRPC5 channel.

Transient receptor potential canonical (TRPC) channels are non-selective calcium-permeable cation channels. Among 7 subunits, TRPC4, 5 channels are well known to be potentiated by direct bindings of calcium ion and phosphatidylinositol 4,5-biphosphate. Both calcium and PIP2 were therefore also implicated in activation and desensitization of the channel. The binding site for calcium ion was revealed but PIP2 binding site remains controversial. Previously, we found two tryptophan residues in TRPC4 channel as a candidate by which calcium sensing in S2-S3 linker is delivered to channel gating. Here, we suggest the WW site in TRPC5 channel conducts delivering of calcium sensing like in TRPC4, and also acts as a potential PIP2 binding site. We made two single mutants (W434A, W435A) and a double mutant (WW/AA) from human TRPC5 channel. Wild-type and mutant TRPC5 channels were expressed in HEK293 cells for macroscopic current recordings in whole-cell or inside-out mode. Generally, our data demonstrate W434 and W435 residues participate in PIP2 binding and calcium sensing. Interestingly, when co-expressed with human muscarinic acetylcholine receptor type 3 (mAChR3) and stimulated by carbachol, W434A channel didn’t show desensitization characteristically observed in wild-type TRPC5 channel. Loss of desensitization in W434A channel implies that it has lost the ability of sensing PIP2 hydrolysis by PLC or the ability of phosphorylation by PKC. The latter possibility is ruled out because the C-terminus of the channels we used were truncated so they do not contain the phosphorylation site. Recently, PIP2-bound ion channel structures were revealed including TRPs. Many channels had PIP2 molecules near S3, S4, and S5 helices. Our electrophysiological data also indicate that PIP2 binds near S3 helix in TRPC5 channel, and the WW site is a specific candidate for sensing PIP2 and calcium.

Development of a high throughput-compatible fluorescence-based assay for TRPM3 channel activity and surface expression.

The Transient Receptor Potential Melastatin 3 (TRPM3) channel is a calcium-permeable non-selective cation channel that can be activated by heat and by a variety of chemical ligands, including neurosteroid pregnenolone sulfate (PS). The TRPM3 channel senses noxious heat in nociceptive neurons, and it is also expressed in the brain where its gating mechanisms and functions remain poorly understood. TRPM3 channel mutations in human patients were recently reported to be associated with intellectual disability and epilepsy, and to cause a gain of function phenotype by increasing the open probability of the channel. By using these mutations along with wild-type TRPM3 channels, expressed in tandem with a genetically encoded fluorescent calcium indicator, we developed and benchmarked a fluorescence-based assay to evaluate ion channel activity. We will apply this fluorescence-based assay to investigate genotype to phenotype relations in the TRPM3 channel at high-throughput. To distinguish between mutations affecting channel activity from those that impact surface expression in the plasma membrane, we are screening diverse engineered TRPM3 channel constructs that would enable specific fluorescence labelling of surface-expressed ion channels. This trafficking assay will further facilitate our effort to systematically identify residues in the transmembrane domain of the channel that are crucial for activation by different types of agonists.

Novel small-molecule modulation of PIEZO1 investigated by conventional and automated patch-clamp.

PIEZO1 trimers form mechanically-activated calcium-permeable non-selective cation channels which are expressed in many types of cells. Mutations in the PIEZO1 gene are associated with generalised lymphatic dysplasia, varicose vein disease, dehydrated hereditary stomatocytosis, malarial resistance and other conditions. Pharmacological modulation of PIEZO1 has been achieved using various compounds, most notably the small-molecule agonist Yoda1 and its inhibitor Dooku1. However, improvements in the potency and pharmacokinetic properties of such compounds are required to increase the usefulness, and small-molecule inhibitors of PIEZO1 are largely lacking. We sought to identify novel small-molecule agonists and inhibitors of mouse and human PIEZO1 channels. Analogues of Yoda1 were generated by medicinal chemistry approaches and a library of 11,200 compounds was screened in a calcium assay using cells overexpressing human PIEZO1. Analogues of hits from the screen were designed and synthesised and then tested for effectiveness in cells overexpressing mouse or human PIEZO1 using an orthogonal calcium assay, conventional (manual) patch-clamp and automated patch-clamp in 384-well format (SyncroPatch 384). We identified Yoda1 analogues with better aqueous solubility and microsomal stability than Yoda1 and improved potency. We identified and validated two novel structurally distinct small-molecule inhibitors of PIEZO1. Automated patch-clamp techniques were refined for activation of PIEZO1 channels by fluid flow in 384-well format. Novel small-molecule modulators of the channels were tested in these automated assays and confirmed as PIEZO1 modulators. We gained preliminary knowledge of the structure-activity relationships and promising compounds for further development. Supported by British Heart Foundation.

Calcium-dependent modulation of BKCa channel activity induced by plasmonic gold nanoparticles in pulmonary artery smooth muscle cells and hippocampal neurons.

Gold nanoparticles are widely used for biomedical applications, but the precise molecular mechanism of their interaction with cellular structures is still unclear. Assuming that intracellular calcium fluctuations associated with surface plasmon-induced calcium entry could modulate the activity of potassium channels, we studied the effect of 5nm gold nanoparticles on calcium-dependent potassium channels and associated calcium signaling in freshly isolated rat pulmonary artery smooth muscle cells and cultured hippocampal neurons. Outward potassium currents were recorded using patch-clamp techniques. Changes in intracellular calcium concentration were measured using the high affinity Ca2+ fluorescent indicator fluo-3 and laser confocal microscope. In pulmonary artery smooth muscle cells, plasmonic gold nanoparticles increased the amplitude of currents via large-conductance Ca2+ -activated potassium channels, which was potentiated by green laser irradiation near plasmon resonance wavelength (532 nm). Buffering of intracellular free calcium with ethylene glycol-bis-N,N,N',N'-tetraacetic acid (EGTA) abolished these effects. Furthermore, using confocal laser microscopy it was found that application of gold nanoparticles caused oscillations of intracellular calcium concentration that were decreasing in amplitude with time. In cultured hippocampal neurons gold nanoparticles inhibited the effect of EGTA slowing down the decline of the BKCa current while partially restoring the amplitude of the slow after hyperpolarizing currents. We conclude that fluctuations in intracellular calcium can modulate plasmonic gold nanoparticles-induced gating of BKCa channels in smooth muscle cells and neurons through an indirect mechanism, probably involving the interaction of plasmon resonance with calcium-permeable ion channels, which leads to a change in intracellular calcium level.

Modulatory Action of Voltage Gated Ion Channels in Inflammation and Inflammatory Pain

Pain is “an unpleasant sensory and emotional experience associated with actual or potential tissue damage, or described by the patient in terms of such damage”. The origin of every pain syndrome is inflammation. A group of voltage-gated channels that are permeable to calcium ions enhances sensory transduction witnessed during inflammation. Hence, understanding calcium signaling is an essential step towards recognizing neural network activity associated with pain management. In this review, we attempted to understand the impact of calcium-permeable ion channels in the recognition, processing, transduction and modulation of pain signals. Results obtained revealed that calcium being one of the most ubiquitous secondary messengers play a significant role in modulating numerous biological processes, including inflammation and pain. Though almost all subtypes of calcium channels are highly expressed in the central nervous system (CNS), the “N-type calcium ion-channels” play an important function at the time of neurotransmitter release from the afferent terminals within the spinal dorsal. Hence, they serve as a key therapeutic target during the treatment of analgesics. Migraine is also reported to involve neurogenic inflammation. “P/Q-type calcium channels” is suggested to have important role in migraine. The inhibition of these channels through various analgesics serves as a treatment against inflammatory and neuropathic pain. However, few of these inhibitors have numerous side effects, including cancer. Hence, these inhibitors may be consumed under the supervision of medical practitioners. In this review, we revealed the understanding and regulation of ion channels in inflammation causing pain and its treatment.

The transient receptor potential melastatin 2: a new therapeutical target for Parkinson's disease?

The transient receptor potential melastatin 2 is a calcium-permeable cation channel member of the TRP family. Also known as an oxidative stress-activated channel, the transient receptor potential melastatin 2 gating mechanism is dependent on reactive oxygen species. In pathological conditions, transient receptor potential melastatin 2 is overactivated, leading to a Ca2+ influx that alters cell homeostasis and promotes cell death. The role of transient receptor potential melastatin 2 in neurodegenerative diseases, including Alzheimer's disease and ischemia, has already been described and reviewed. However, data on transient receptor potential melastatin 2 involvement in Parkinson's disease pathology has emerged only in recent years and the issue lacks review studies that focus specifically on this topic. The present review aims to elucidate the role of the transient receptor potential melastatin 2 channel in Parkinson's disease by reviewing, summarizing, and discussing the in vitro, in vivo, and human studies published until August 2022. Here we describe fourteen studies that evaluated the transient receptor potential melastatin 2 channel in Parkinson's disease. The Parkinson's disease model used, transient receptor potential melastatin 2 antagonist and genetic approaches, and the main outcomes reported were discussed. The studies described transient receptor potential melastatin 2 activation and enhanced expression in different Parkinson's disease models. They also evidenced protective and restorative effects when using transient receptor potential melastatin 2 antagonists, knockout, or silencing. This review provides a literature overview and suggests where there is a need for more research. As a perspective point, this review shows evidence that supports transient receptor potential melastatin 2 as a pharmacological target for Parkinson's disease in the future.

Calcium Channels, OST1 and Stomatal Defence: Current Status and Beyond

Stomatal immunity is regulated by pathogen-associated molecular patterns (PAMPs)- and abscisic acid (ABA)-triggered signalling in different ways. Cytoplasmic Ca2+ signature in the guard cells plays a vital function in stomatal immunity, but the mechanism of Ca2+ import is unknown. It has been very recently established that the hyperosmolality-gated calcium-permeable channels (OSCAs) and cyclic nucleotide-gated channels (CNGCs) are responsible for the influx of Ca2+ in the cytoplasm, which are activated after BIK1-mediated phosphorylation and ABA interaction during PAMPs- and ABA-triggered stomatal immunity in plants, respectively. Further, ABA-triggered OPEN STOMATA1 (OST1) causes the disassembly of microtubules in the guard cells besides activation of S-type anion channels (SLAC1) for the efflux of cytoplasmic anions that leads to stomata closure.

Genome-wide analysis of hyperosmolality-gated calcium-permeable channel (OSCA) family members and their involvement in various osmotic stresses in Brassica napus

Plant hyperosmolality-gated calcium-permeable channel (OSCA) is a calcium permeable cation channel that responds to hyperosmotic stress and plays a pivotal role in plant growth, development and stress response. Through a genome-wide survey, 41 OSCA genes were identified from the genome of Brassica napus. The OSCA family genes were unevenly distributed over 14 chromosomes of B. napus and phylogenetic analysis separated the OSCA family into four clades. Motif analyses indicated that OSCA proteins in the same clade were highly conserved and the protein conserved motifs shared similar composition patterns. The OSCA promoter regions contained many hormone-related elements and stress response elements. Gene duplication analysis elucidated that WGD/segmental duplication was the main driving force for the expansion of OSCA genes during evolution and these genes mainly underwent purifying selection. RNA-seq and qRT-PCR analysis of different tissues showed that OSCA genes are expressed and function mainly in the root. Among these genes, BnOSCA3.1a and BnOSCA3.1c had relatively high expression levels under osmotic stresses and cold stress and were highly expressed in different tissues. Protein interaction network analysis showed that a total of 5802 proteins might interact with OSCAs in B. napus, while KEGG/GO enrichment analysis indicated that OSCAs and their interacting proteins were mainly involved in plant response to abiotic stress. This systematic analysis of the OSCAs in B. napus identified gene structures, evolutionary features, expression patterns and related biological processes. These findings will facilitate further functional and evolutionary analysis of OSCAs in B. napus for breeding of osmotic-stress-resistant plants.

The eINTACT system dissects bacterial exploitation of plant osmosignalling to enhance virulence

Bacteria inject effector proteins into host cells to manipulate cellular processes that promote disease. Since bacteria deliver minuscule amounts of effectors only into targeted host cells, it is technically challenging to capture effector-dependent cellular changes from bulk-infected host tissues. Here, we report a new technique called effector-inducible isolation of nuclei tagged in specific cell types (eINTACT), which facilitates affinity-based purification of nuclei from Arabidopsis plant cells that have received Xanthomonas bacterial effectors. Analysis of purified nuclei reveals that the Xanthomonas effector XopD manipulates the expression of Arabidopsis abscisic acid signalling-related genes and activates OSCA1.1, a gene encoding a calcium-permeable channel required for stomatal closure in response to osmotic stress. The loss of OSCA1.1 causes leaf wilting and reduced bacterial growth in infected leaves, suggesting that OSCA1.1 promotes host susceptibility. eINTACT allows us to uncover that XopD exploits host OSCA1.1/abscisic acid osmosignalling-mediated stomatal closure to create a humid habitat that favours bacterial growth and opens up a new avenue for accurately elucidating functions of effectors from numerous gram-negative plant bacteria in native infection contexts.

Co-chaperones of the Human Endoplasmic Reticulum: An Update.

In mammalian cells, the rough endoplasmic reticulum (ER) plays central roles in the biogenesis of extracellular plus organellar proteins and in various signal transduction pathways. For these reasons, the ER comprises molecular chaperones, which are involved in import, folding, assembly, export, plus degradation of polypeptides, and signal transduction components, such as calcium channels, calcium pumps, and UPR transducers plus adenine nucleotide carriers/exchangers in the ER membrane. The calcium- and ATP-dependent ER lumenal Hsp70, termed immunoglobulin heavy-chain-binding protein or BiP, is the central player in all these activities and involves up to nine different Hsp40-type co-chaperones, i.e., ER membrane integrated as well as ER lumenal J-domain proteins, termed ERj or ERdj proteins, two nucleotide exchange factors or NEFs (Grp170 and Sil1), and NEF-antagonists, such as MANF. Here we summarize the current knowledge on the ER-resident BiP/ERj chaperone network and focus on the interaction of BiP with the polypeptide-conducting and calcium-permeable Sec61 channel of the ER membrane as an example for BiP action and how its functional cycle is linked to ER protein import and various calcium-dependent signal transduction pathways.