Ca2+-dependent Protein Kinase Research Articles

Calcium signaling facilitates chilling- and GA- induced dormancy release in tree peony.

Calcium plays a crucial role in plant growth and development, yet little is known about its function in endodormancy regulation. Tree peony (Paeonia suffruticosa), characterized by compound buds and large flowers, is well-known for its ornamental and medicinal value. To break bud dormancy release is a prerequisite of flowering and forcing culture, particularly during the Spring Festival. In this study, the Ca2+ chelator EGTA and Ca2+ channel blocker LaCl3 were applied, resulting in a significant delay in budburst during both chilling- and gibberellin (GA)- induced dormancy release in a dosage-dependent manner. As expected, the retardation of bud break was recovered by the supplementation of 30 mM CaCl2, indicating a facilitating role of calcium in dormancy release. Accordingly, several calcium-sensor-encoding genes including Calmodulin (CaM) and Ca2+-dependent protein kinases (CDPKs) were significantly up-regulated by prolonged chilling and exogenous GAs. Ultrastructure observations revealed a decline in starch grains and the reopening of transport corridors following prolonged chilling. Calcium deposits were abundant in the cell walls and intercellular spaces at the early dormant stage but were enriched in the cytosol and nucleus before dormancy release. Additionally, several genes associated with dormancy release, including EBB1, EBB3, SVP, GA20ox, RGL1, BG6, and BG9, were differentially expressed after calcium blocking and recovery treatments, indicating that calcium might partially modulate dormancy release through GA and ABA pathways. Our findings provide novel insights into the mechanism of dormancy release and offer potential benefits for improving and perfecting forcing culture technology in tree peonies.

In Vivo Phosphorylation of the Cytosolic Glucose-6-Phosphate Dehydrogenase Isozyme G6PD6 in Phosphate-Resupplied Arabidopsis thaliana Suspension Cells and Seedlings.

Glucose-6-phosphate dehydrogenase (G6PD) catalyzes the first committed step of the oxidative pentose phosphate pathway (OPPP). Our recent phosphoproteomics study revealed that the cytosolic G6PD6 isozyme became hyperphosphorylated at Ser12, Thr13 and Ser18, 48 h following phosphate (Pi) resupply to Pi-starved (-Pi) Arabidopsis thaliana cell cultures. The aim of the present study was to assess whether G6PD6 phosphorylation also occurs in shoots or roots following Pi resupply to -Pi Arabidopsis seedlings, and to investigate its relationship with G6PD activity. Interrogation of phosphoproteomic databases indicated that N-terminal, multi-site phosphorylation of G6PD6 and its orthologs is quite prevalent. However, the functions of these phosphorylation events remain unknown. Immunoblotting with an anti-(pSer18 phosphosite-specific G6PD6) antibody confirmed that G6PD6 from Pi-resupplied, but not -Pi, Arabidopsis cell cultures or seedlings (i.e., roots) was phosphorylated at Ser18; this correlated with a significant increase in extractable G6PD activity, and biomass accumulation. Peptide kinase assays of Pi-resupplied cell culture extracts indicated that G6PD6 phosphorylation at Ser18 is catalyzed by a Ca2+-dependent protein kinase (CDPK), which correlates with the 'CDPK-like' targeting motif that flanks Ser18. Our results support the hypothesis that N-terminal phosphorylation activates G6PD6 to enhance OPPP flux and thus the production of reducing power (i.e., NADPH) and C-skeletons needed to establish the rapid resumption of growth that ensures Pi-resupply to -Pi Arabidopsis.

Overexpression of AcCDPK1 and AcCDPK5 from Allium cepa. L enhances Phytophthora nicotianae resistance in Nicotiana benthamiana

The Ca2+-dependent protein kinases (CDPKs) play important roles in Ca2+ signal transduction pathway in plant biotic stress response. In this study, AcCDPK1 and AcCDPK5 were isolated from onion. AcCDPK1 and AcCDPK5 showed higher expression in Phytophthora nicotianae resistant onion genotype compared to the susceptible onion line. AcCDPK1 and AcCDPK5 were cell membrane-localized proteins and contained conserved regions of plant CDPK protein, such as a serine‑ threonine protein kinase domain and a C-terminal calmodulin-like domain. AcCDPK1 and AcCDPK5 ectopic overexpression in Nicotiana benthamiana presented milder disease symptoms than that of wild type after P. nicotianae inoculation. The overexpression of AcCDPK1 and AcCDPK5 promoted the accumulation of ROS and upregulated the NbRBOHD transcript level at the initial stage of inoculation. In addition, AcCDPK1 and AcCDPK5 overexpression promoted the expression of NbPR1, which was related to the pathogen resistance. AcCDPK1 performed more obvious roles than AcCDPK5 in P. nicotianae resistance. The results suggested that AcCDPK1 and AcCDPK5 played a positive role in P. nicotianae resistance by regulating ROS accumulation and downstream resistance related genes expression.

Benzotriazole Ultraviolet Stabilizers (BUVSs) as Potential Protein Kinase Antagonists in Rice.

The ubiquitous occurrence of benzotriazole ultraviolet stabilizers (BUVSs) in the environment and organisms has warned of their potential ecological and health risks. Studies showed that some BUVSs exerted immune and chronic toxicities to animals by disturbing signaling transduction, yet limited research has investigated the toxic effects on crop plants and the underlying mechanisms of signaling regulation. Herein, a laboratory-controlled hydroponic experiment was conducted on rice to explore the phytotoxicity of BUVSs by integrating conventional biochemical experiments, transcriptomic analysis, competitive sorption assays, and computational studies. The results showed that BUVSs inhibited the growth of rice by 6.30-20.4% by excessively opening the leaf stomas, resulting in increased transpiration. BUVSs interrupted the transduction of abscisic acid (ABA) signal through competitively binding to Ca2+-dependent protein kinase (CDPK), weakening the CDPK phosphorylation and further inhibiting the downstream signaling. As structural analogues of ATP, BUVSs acted as potential ABA signaling antagonists, leading to physiological dysfunction in mediating stomatal closure under stresses. This is the first comprehensive study elucidating the effects of BUVSs on the function of key proteins and the associated signaling transduction in plants and providing insightful information for the risk evaluation and control of BUVSs.

Analysis of CDPK1 targets identifies a trafficking adaptor complex that regulates microneme exocytosis in Toxoplasma.

Apicomplexan parasites use Ca2+-regulated exocytosis to secrete essential virulence factors from specialized organelles called micronemes. Ca2+-dependent protein kinases (CDPKs) are required for microneme exocytosis; however, the molecular events that regulate trafficking and fusion of micronemes with the plasma membrane remain unresolved. Here, we combine sub-minute resolution phosphoproteomics and bio-orthogonal labeling of kinase substrates in Toxoplasma gondii to identify 163 proteins phosphorylated in a CDPK1-dependent manner. In addition to known regulators of secretion, we identify uncharacterized targets with predicted functions across signaling, gene expression, trafficking, metabolism, and ion homeostasis. One of the CDPK1 targets is a putative HOOK activating adaptor. In other eukaryotes, HOOK homologs form the FHF complex with FTS and FHIP to activate dynein-mediated trafficking of endosomes along microtubules. We show the FHF complex is partially conserved in T. gondii, consisting of HOOK, an FTS homolog, and two parasite-specific proteins (TGGT1_306920 and TGGT1_316650). CDPK1 kinase activity and HOOK are required for the rapid apical trafficking of micronemes as parasites initiate motility. Moreover, parasites lacking HOOK or FTS display impaired microneme protein secretion, leading to a block in the invasion of host cells. Taken together, our work provides a comprehensive catalog of CDPK1 targets and reveals how vesicular trafficking has been tuned to support a parasitic lifestyle.

Analysis of CDPK1 targets identifies a trafficking adaptor complex that regulates microneme exocytosis in Toxoplasma

Apicomplexan parasites use Ca2+-regulated exocytosis to secrete essential virulence factors from specialized organelles called micronemes. Ca2+-dependent protein kinases (CDPKs) are required for microneme exocytosis; however, the molecular events that regulate trafficking and fusion of micronemes with the plasma membrane remain unresolved. Here, we combine sub-minute resolution phosphoproteomics and bio-orthogonal labeling of kinase substrates in Toxoplasma gondii to identify 163 proteins phosphorylated in a CDPK1-dependent manner. In addition to known regulators of secretion, we identify uncharacterized targets with predicted functions across signaling, gene expression, trafficking, metabolism, and ion homeostasis. One of the CDPK1 targets is a putative HOOK activating adaptor. In other eukaryotes, HOOK homologs form the FHF complex with FTS and FHIP to activate dynein-mediated trafficking of endosomes along microtubules. We show the FHF complex is partially conserved in T. gondii, consisting of HOOK, an FTS homolog, and two parasite-specific proteins (TGGT1_306920 and TGGT1_316650). CDPK1 kinase activity and HOOK are required for the rapid apical trafficking of micronemes as parasites initiate motility. Moreover, parasites lacking HOOK or FTS display impaired microneme protein secretion, leading to a block in the invasion of host cells. Taken together, our work provides a comprehensive catalog of CDPK1 targets and reveals how vesicular trafficking has been tuned to support a parasitic lifestyle.

NILR1 perceives a nematode ascaroside triggering immune signaling and resistance

Plants use pattern recognition receptors (PRRs) to perceive conserved molecular patterns derived from pathogens and pests, thereby activating a sequential set of rapid cellular immune responses, including activation of mitogen-activated protein kinases (MAPKs) and Ca2+-dependent protein kinases (CDPKs), transcriptional reprogramming (particularly the induction of defense-related genes), ion fluxes, and production of reactive oxygen species.1 Plant PRRs belong to the multi-membered protein families of receptor-like kinases (RLKs) or receptor-like proteins (RLPs). RLKs consist of a ligand-binding ectodomain, a single-pass transmembrane domain, and an intracellular kinase domain, while RLPs possess the same functional domains, except for the intracellular kinase domain.2 The most abundant nematode ascaroside, Ascr18, is a nematode-associated molecular pattern (NAMP) that induces immune signaling and enhances resistance to pathogens and pests in various plant species.3 In this study, we found that the Arabidopsis NEMATODE-INDUCED LRR-RLK1 (NILR1) protein4 physically interacts with the Ascr18 elicitor, as indicated by a specific direct interaction between NILR1 and Ascr18, and NILR1 is genetically required for Ascr18-triggered immune signaling and resistance to both bacterium and nematode, as manifested by the abolishment of these immune responses in the nilr1 mutant. These results suggest that NILR1 is the immune receptor of the nematode NAMP Ascr18, mediating Ascr18-triggered immune signaling and resistance to pathogens and pests.

Substrate profiling of the Arabidopsis Ca2+-dependent protein kinase AtCPK4 and its Ricinus communis ortholog RcCDPK1

AtCPK4 and AtCPK11 are Arabidopsis thaliana Ca2+-dependent protein kinase (CDPK) paralogs that have been reported to positively regulate abscisic acid (ABA) signal transduction by phosphorylating ABA-responsive transcription factor-4 (AtABF4). By contrast, RcCDPK1, their closest Ricinus communis ortholog, participates in the control of anaplerotic carbon flux in developing castor oil seeds by catalyzing inhibitory phosphorylation of bacterial-type phosphoenolpyruvate carboxylase at Ser451. LC-MS/MS revealed that AtCPK4 and RcCDPK1 transphosphorylated several common, conserved residues of AtABF4 and its castor ortholog, TRANSCRIPTION FACTOR RESPONSIBLE FOR ABA REGULATON. Arabidopsis atcpk4/atcpk11 mutants displayed an ABA-insensitive phenotype that corroborated the involvement of AtCPK4/11 in ABA signaling. A kinase-client assay was employed to identify additional AtCPK4/RcCDPK1 targets. Both CDPKs were separately incubated with a library of 2095 peptides representative of Arabidopsis protein phosphosites; five overlapping targets were identified including PLANT INTRACELLULAR RAS-GROUP-RELATED LEUCINE-RICH REPEAT PROTEIN-9 (AtPIRL9) and the E3-ubiquitin ligase ARABIDOPSIS TOXICOS EN LEVADURA 6 (AtATL6). AtPIRL9 and AtATL6 residues phosphorylated by AtCPK4/RcCDPK1 conformed to a CDPK recognition motif that was conserved amongst their respective orthologs. Collectively, this study provides evidence for novel AtCPK4/RcCDPK1 substrates, which may help to expand regulatory networks linked to Ca2+- and ABA-signaling, immune responses, and central carbon metabolism.

Delineating Calcium Signaling Machinery in Plants: Tapping the Potential through Functional Genomics.

Plants have developed calcium (Ca2+) signaling as an important mechanism of regulation of stress perception, developmental cues, and responsive gene expression. The post-genomic era has witnessed the successful unravelling of the functional characterization of genes and the creation of large datasets of molecular information. The major elements of Ca2+ signaling machinery include Ca2+ sensors and responders such as Calmodulins (CaMs), Calmodulin-like proteins (CMLs), Ca2+/CaM-dependent protein kinases (CCaMKs), Ca2+-dependent protein kinases (CDPKs), Calcineurin B-like proteins (CBLs) as well as transporters, such as Cyclic nucleotide-gated channels (CNGCs), Glutamate-like receptors (GLRs), Ca2+-ATPases, Ca2+/H+ exchangers (CAXs) and mechanosensitive channels. These elements play an important role in the regulation of physiological processes and plant responses to various stresses. Detailed genomic analysis can help us in the identification of potential molecular targets that can be exploited towards the development of stress-tolerant crops. The information sourced from model systems through omics approaches helps in the prediction and simulation of regulatory networks involved in responses to different stimuli at the molecular and cellular levels. The molecular delineation of Ca2+ signaling pathways could be a stepping stone for engineering climate-resilient crop plants. Here, we review the recent developments in Ca2+ signaling in the context of transport, responses, and adaptations significant for crop improvement through functional genomics approaches.

TypiCal but DeliCate Ca++re: Dissecting the Essence of Calcium Signaling Network as a Robust Response Coordinator of Versatile Abiotic and Biotic Stimuli in Plants.

Plant growth, development, and ultimately crop productivity are largely impacted by the interaction of plants with different abiotic and biotic factors throughout their life cycle. Perception of different abiotic stresses, such as salt, cold, drought, heat, and heavy metals, and interaction with beneficial and harmful biotic agents by plants lead to transient, sustained, or oscillatory changes of [calcium ion, Ca2+]cyt within the cell. Significant progress has been made in the decoding of Ca2+ signatures into downstream responses to modulate differential developmental and physiological responses in the whole plant. Ca2+ sensor proteins, mainly calmodulins (CaMs), calmodulin-like proteins (CMLs), and others, such as Ca2+-dependent protein kinases (CDPKs), calcineurin B-like proteins (CBLs), and calmodulin-binding transcription activators (CAMTAs) have played critical roles in coupling the specific stress stimulus with an appropriate response. This review summarizes the current understanding of the Ca2+ influx and efflux system in plant cells and various Ca2+ binding protein-mediated signal transduction pathways that are delicately orchestrated to mitigate abiotic and biotic stresses. The probable interactions of different components of Ca2+ sensor relays and Ca2+ sensor responders in response to various external stimuli have been described diagrammatically focusing on established pathways and latest developments. Present comprehensive insight into key components of the Ca2+ signaling toolkit in plants can provide an innovative framework for biotechnological manipulations toward crop improvability in near future.

Truncated variants of Ca2+-dependent protein kinases: a conserved regulatory mechanism?

Recent studies suggest that immune-induced alternative splice variants of the Arabidopsis thaliana Ca2+-dependent protein kinase (CDPK) AtCPK28 may result in signal attenuation. We put forward the hypothesis that expression of alternative truncated variants may be a broadly conserved regulatory mechanism of CDPKs throughout the green lineage.

Molecular Mechanism of Stimulation of Na-K-ATPase by Leukotriene D4 in Intestinal Epithelial Cells.

Na-K-ATPase provides a favorable transcellular Na gradient required for the functioning of Na-dependent nutrient transporters in intestinal epithelial cells. The primary metabolite for enterocytes is glutamine, which is absorbed via Na-glutamine co-transporter (SN2; SLC38A5) in intestinal crypt cells. SN2 activity is stimulated during chronic intestinal inflammation, at least in part, secondarily to the stimulation of Na-K-ATPase activity. Leukotriene D4 (LTD4) is known to be elevated in the mucosa during chronic enteritis, but the way in which it may regulate Na-K-ATPase is not known. In an in vitro model of rat intestinal epithelial cells (IEC-18), Na-K-ATPase activity was significantly stimulated by LTD4. As LTD4 mediates its action via Ca-dependent protein kinase C (PKC), Ca levels were measured and were found to be increased. Phorbol 12-myristate 13-acetate (PMA), an activator of PKC, also mediated stimulation of Na-K-ATPase like LTD4, while BAPTA-AM (Ca chelator) and calphostin-C (Cal-C; PKC inhibitor) prevented the stimulation of Na-K-ATPase activity. LTD4 caused a significant increase in mRNA and plasma membrane protein expression of Na-K-ATPase α1 and β1 subunits, which was prevented by calphostin-C. These data demonstrate that LTD4 stimulates Na-K-ATPase in intestinal crypt cells secondarily to the transcriptional increase of Na-K-ATPase α1 and β1 subunits, mediated via the Ca-activated PKC pathway.

Transcriptional regulation and stress-defensive key genes induced by γ-aminobutyric acid in association with tolerance to water stress in creeping bentgrass

ABSTRACT γ-Aminobutyric acid (GABA) acts as an important regulator involved in the mediation of cell signal transduction and stress tolerance in plants. However, the function of GABA in transcriptional regulation is not fully understood in plants under water stress. The creeping bentgrass (Agrostis stolonifera) was pretreated with or without GABA (0.5 mM) for 24 hours before being exposed to 5 days of water stress. Physiological analysis showed that GABA-treated plants maintained significantly higher endogenous GABA content, leaf relative water content, net photosynthetic rate, and lower osmotic potential than untreated plants under water stress. The GABA application also significantly alleviated stress-induced increases in superoxide anion (O2 .-) content, hydrogen peroxide (H2O2) content, and electrolyte leakage through enhancing total antioxidant capacity, superoxide dismutase (SOD) activity, and peroxidase (POD) activity in response to water stress. The transcriptomic analysis demonstrated that the GABA-induced changes in differentially expressed genes (DEGs) involved in carbohydrates, amino acids, and secondary metabolism helped to maintain better osmotic adjustment, energy supply, and metabolic homeostasis when creeping bentgrass suffers from water stress. The GABA triggered Ca2+-dependent protein kinase (CDPK) signaling and improved transcript levels of DREB1/2 and WRKY1/24/41 that could be associated with the upregulation of stress-related functional genes such as POD, DHNs, and HSP70 largely contributing to improved tolerance to water stress in relation to the antioxidant, prevention of cell dehydration, and protein protection in leaves.

Seed priming with H2S and Ca2+ trigger signal memory that induces cross-adaptation against nickel stress in zucchini seedlings

In this study, the effect of seed priming with sodium hydro sulfide (NaHS) and CaCl2 as well as the possible relationship between them in inducing post-germinative cross-adaptation in zucchini seedlings (cv Courgette d’Italie) were investigated. Results showed that Ni toxicity reduced plant growth and photosynthetic pigments, decreased the content of ascorbate (AsA) and total thiols, increased hydrogen peroxide (H2O2) content and electrolyte leakage (EL), up-regulated the transcription levels of Ca2+-dependent protein kinase (CDPK) and phytochelatin (PCs) genes and elevated H2S content in leaves of zucchini seedlings. Individual or combined seed priming with Ca2+ and NaHS improved the content of photosynthetic pigments and seedling biomass, reduced H2O2 content and EL, increased the content of AsA and total thiols, decreased ascorbate peroxidase activity and enhanced glutathione reductase activity in leaves. These findings suggest the last time effect of seed priming with Ca2+ and NaHS on inducing cross-adaptation in seedlings under Ni stress. H2S accumulation and other responses induced with Ca2+ in leaves were weakened with hypotaurine (HT as H2S scavenger), denoting seed priming with Ca2+ established cross-adaptation in a H2S-dependent manner. Seed priming with NaHS amplified CDPK transcripts in leaves of seedlings and seed priming with ethylene glycol tetraacetic acid (as Ca2+ chelator), lanthanum chloride and verapamil (as plasma membrane channel blockers) reduced transcript levels of CDPK and PCs genes and reversed impacts of seed priming with NaHS. These results indicated that the cross-adaptation induced with NaHS is mediated through Ca2+ signaling. Overall our findings suggest that two-side cross-talk between Ca2+ and H2S is involved in the acquisition of a signal memory in seed embryo cells which can be employed upon a later Ni-exposure and more strongly enhance AsA-GSH cycle, redox homeostasis and phytochelatin transcripts in leaves of zucchini seedlings grown from primed seeds.

Involvement of phosphatidylinositol metabolism in aluminum-induced malate secretion in Arabidopsis.

To identify the upstream signaling of aluminum-induced malate secretion through aluminum-activated malate transporter 1 (AtALMT1), a pharmacological assay using inhibitors of human signal transduction pathways was performed. Early aluminum-induced transcription of AtALMT1 and other aluminum-responsive genes was significantly suppressed by phosphatidylinositol 4-kinase (PI4K) and phospholipase C (PLC) inhibitors, indicating that the PI4K-PLC metabolic pathway activates early aluminum signaling. Inhibitors of phosphatidylinositol 3-kinase (PI3K) and PI4K reduced aluminum-activated malate transport by AtALMT1, suggesting that both the PI3K and PI4K metabolic pathways regulate this process. These results were validated using T-DNA insertion mutants of PI4K and PI3K-RNAi lines. A human protein kinase inhibitor, putatively inhibiting homologous calcineurin B-like protein-interacting protein kinase and/or Ca-dependent protein kinase in Arabidopsis, suppressed late-phase aluminum-induced expression of AtALMT1, which was concomitant with the induction of an AtALMT1 repressor, WRKY46, and suppression of an AtALMT1 activator, Calmodulin-binding transcription activator 2 (CAMTA2). In addition, a human deubiquitinase inhibitor suppressed aluminum-activated malate transport, suggesting that deubiquitinases can regulate this process. We also found a reduction of aluminum-induced citrate secretion in tobacco by applying inhibitors of PI3K and PI4K. Taken together, our results indicated that phosphatidylinositol metabolism regulates organic acid secretion in plants under aluminum stress.

Crosstalk During Fruit Ripening and Stress Response Among Abscisic Acid, Calcium-Dependent Protein Kinase and Phenylpropanoid

Phenylpropanoids are secondary metabolites produced by plants. They, by differential expression, are involved in responses to biotic and abiotic stresses and confer plant plasticity. In addition, they are synthesized under normal conditions during the fruit-ripening process. Therefore, the understanding of the mechanics involved in the accumulation of these compounds in plants is of extreme importance for the development of plants with greater resistance and tolerance to biotic and abiotic stresses, and plants with greater functional potential. There is evidence that one of the pathways of the induction of phenylpropanoids is dependent on abscisic acid (ABA) and it is generated by a signaling cascade involving calcium (Ca2+) and Ca2+-dependent protein kinases (CDPKs). Plants have several Ca2+ binding proteins that act as cellular sensors and represent the first points of signal transduction. CDPKs are mono-molecular Ca2+-sensor/kinase-effector proteins, which perceive Ca2+ signals and translate them into protein phosphorylation and thus represent an ideal tool for signal transduction. However, the mechanisms involved in the ABA–CDPK–phenylpropanoids crosstalk under stress conditions and during fruit ripening remains uncertain. Therefore, this review seeks to surface a new line of evidence as an attempt to understand the manner in which the induction of phenylpropanoids occurs in plants.

The Arabidopsis Ca2+-Dependent Protein Kinase CPK12 Is Involved in Plant Response to Salt Stress.

CDPKs (Ca2+-Dependent Protein Kinases) are very important regulators in plant response to abiotic stress. The molecular regulatory mechanism of CDPKs involved in salt stress tolerance remains unclear, although some CDPKs have been identified in salt-stress signaling. Here, we investigated the function of an Arabidopsis CDPK, CPK12, in salt-stress signaling. The CPK12-RNA interference (RNAi) mutant was much more sensitive to salt stress than the wild-type plant GL1 in terms of seedling growth. Under NaCl treatment, Na+ levels in the roots of CPK12-RNAi plants increased and were higher than levels in GL1 plants. In addition, the level of salt-elicited H2O2 production was higher in CPK12-RNAi mutants than in wild-type GL1 plants after NaCl treatment. Collectively, our results suggest that CPK12 is required for plant adaptation to salt stress.

Regulation of Plant Immune Signaling by Calcium-Dependent Protein Kinases.

Activation of Ca2+ signaling is a universal response to stress that allows cells to quickly respond to environmental cues. Fluctuations in cytosolic Ca2+ are decoded in plants by Ca2+-sensing proteins such as Ca2+-dependent protein kinases (CDPKs). The perception of microbes results in an influx of Ca2+ that activates numerous CDPKs responsible for propagating immune signals required for resistance against disease-causing pathogens. This review describes our current understanding of CDPK activation and regulation, and provides a comprehensive overview of CDPK-mediated immune signaling through interaction with various substrates.

Principles of Calcium Signal Generation and Transduction in Plant Cells

Calcium ions exhibit unique properties and a universal ability to transmit diverse signals in plant cells under the primary action of hormones, pathogens, light, gravity, and various abiotic stressors. In the last few years, considerable progress has been achieved in deciphering the mechanisms of Ca2+ involvement in the regulation of plant responses. Recent studies revealed the genes encoding Ca2+-permeable channels that conduct Ca2+ currents across the membranes during the transduction of the Ca2+ signal. These proteins comprise the ligand-gated Ca2+-permeable channels activated by cyclic nucleotides (CNGC) and amino acids (glutamate receptor-like channels, GLR), the voltage-gated tonoplast channel (two-pore channel, TPC1), mechanosensitive channels (MSL, MCA, OSCA1), and annexins. The role of Ca2+-ATPase and Ca2+/H+-exchangers in the active extrusion of excess cytoplasmic Ca2+ into the apoplast or cell organelles was examined in detail. The calmodulins (CaM), CaM-like proteins (CML), Ca2+-dependent protein kinases (CDPK), and complexes of calcineurin-B-like proteins (CBL) with CBL-interacting protein kinases (CIPK) were found to produce intricate signaling networks that decode Ca2+ signals and elicit plant responses to external stimuli. This review analyzes the data accumulated over the past decade on the principles of formation and propagation of the calcium signal in plant cells.

Comparative proteomic analysis of Ulva prolifera response to high temperature stress

BackgroundUlva prolifera belongs to green macroalgae and is the dominant species of green tide. It is distributed worldwide and is therefore subject to high-temperature stress during the growth process. However, the adaptation mechanisms of the response of U. prolifera to high temperatures have not been clearly investigated yet.MethodsIn this study, isobaric tags for relative and absolute quantitation (iTRAQ) labelling was applied in combination with the liquid chromatography-tandem mass spectrometry (LC-MS/MS) to conduct comparative proteomic analysis of the response of U. prolifera to high-temperature stress and to elucidate the involvement of this response in adaptation mechanisms. Differentially expressed proteins (DEPs) of U. prolifera under high temperature (denote UpHT) compared with the control (UpC) were identified. Bioinformatic analyses including GO analysis, pathway analysis, and pathway enrichment analysis was performed to analyse the key metabolic pathways that underlie the thermal tolerance mechanism through protein networks. Quantitative real-time PCR and western blot were performed to validate selected proteins.ResultsIn the present study, 1223 DEPs were identified under high temperature compared with the control, which included 790 up-regulated and 433 down-regulated proteins. The high-temperature stimulus mainly induced the expression of glutathione S-transferase, heat shock protein, ascorbate peroxidase, manganese superoxide dismutase, ubiquitin-related protein, lhcSR, rubisco activase, serine/threonine protein kinase 2, adenylate kinase, Ca2+-dependent protein kinase (CDPK), disease resistance protein EDS1, metacaspase type II, NDPK2a, 26S proteasome regulatory subunit, ubiquinone oxidoreductase, ATP synthase subunit, SnRK2s, and cytochrome P450. The down-regulated proteins were photosynthesis-related proteins, glutathione reductase, catalase-peroxidase, thioredoxin, thioredoxin peroxidase, PP2C, and carbon fixation-related proteins. Furthermore, biological index analysis indicated that protein content and SOD activity decreased; the value of Fv/Fm dropped to the lowest point after culture for 96 h. However, APX activity and MDA content increased under high temperature.ConclusionThe present study implied an increase in proteins that were associated with the stress response, oxidative phosphorylation, the cytokinin signal transduction pathway, the abscisic acid signal transduction pathway, and the glutathione metabolism pathway. Proteins that were associated with photosynthesis, carbon fixation in photosynthesis organisms, and the photosynthesis antenna protein pathway were decreased. These pathways played a pivotal role in high temperature regulation. These novel proteins provide a good starting point for further research into their functions using genetic or other approaches. These findings significantly improve the understanding of the molecular mechanisms involved in the tolerance of algae to high-temperature stress.