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Summary: CaWRKY40 in pepper (Capsicum annuum) promotes immune responses to Ralstonia solanacearum infection (RSI) and to high‐temperature, high‐humidity (HTHH) stress, but how it interacts with upstream signalling components remains poorly understood.Here, using approaches of reverse genetics, biochemical and molecular biology we functionally characterised the relationships among the WRKYGMK‐containing WRKY protein CaWRKY27b, the calcium‐dependent protein kinase CaCDPK29, and CaWRKY40 during pepper response to RSI or HTHH.Our data indicate that CaWRKY27b is upregulated and translocated from the cytoplasm to the nucleus upon phosphorylation of Ser137 in the nuclear localisation signal by CaCDPK29. Using electrophoretic mobility shift assays and microscale thermophoresis, we observed that, due to the replacement of Q by M in the conserved WRKYGQK, CaWRKY27b in the nucleus failed to bind to W‐boxes in the promoters of immunity‐ and thermotolerance‐related marker genes. Instead, CaWRKY27b interacted with CaWRKY40 and promoted its binding and positive regulation of the tested marker genes including CaNPR1, CaDEF1 and CaHSP24. Notably, mutation of the WRKYGMK motif in CaWRKY27b to WRKYGQK restored the W‐box binding ability.Our data therefore suggest that CaWRKY27b is phosphorylated by CaCDPK29 and acts as a transcriptional activator of CaWRKY40 during the pepper response to RSI and HTHH. [ABSTRACT FROM AUTHOR]

Malus baccata (L.) Borkh. is widely used as an apple rootstock in northern China. Previous studies have shown that jasmonic acid (JA) can alleviate the oxidative damage in M. baccata roots under low root-zone temperature (LT), yet the regulatory mechanism is unclear. To uncover the mechanism of JA in regulating the response of roots to LT, the differential expression of proteins was profiled using TMT combined with MRM. Of the 4979 proteins quantified, 403 differentially expressed proteins (DEPs) responded to the LT treatment and 264 DEPs responded to the JA + LT treatment. A bioinformatic analysis showed that protein processing in the endoplasmic reticulum, alpha-linolenic acid metabolism, and secondary metabolites play key roles in the regulation of JA in M. baccata roots exposed to LT. In addition, proteins involved in the stress response, such as peroxidases and calcium-dependent protein kinase, also participated in the regulation of JA. The quantitative data were then confirmed using MRM, thereby proving that the TMT results were reliable. The results can provide theoretical reference for the regulation of apple roots under low-temperature stress. [ABSTRACT FROM AUTHOR]

Transmission of malaria-causing parasites to mosquitoes relies on the production of gametocyte stages and their development into gametes. These stages display various microtubule cytoskeletons and the architecture of the corresponding microtubule organisation centres (MTOC) remains elusive. Combining ultrastructure expansion microscopy (U-ExM) with bulk proteome labelling, we first reconstructed in 3D the subpellicular microtubule network which confers cell rigidity to Plasmodium falciparum gametocytes. Upon activation, as the microgametocyte undergoes three rounds of endomitosis, it also assembles axonemes to form eight flagellated microgametes. U-ExM combined with Pan-ExM further revealed the molecular architecture of the bipartite MTOC coordinating mitosis with axoneme formation. This MTOC spans the nuclear membrane linking cytoplasmic basal bodies to intranuclear bodies by proteinaceous filaments. In P. berghei, the eight basal bodies are concomitantly de novo assembled in a SAS6- and SAS4-dependent manner from a deuterosome-like structure, where centrin, γ-tubulin, SAS4 and SAS6 form distinct subdomains. Basal bodies display a fusion of the proximal and central cores where centrin and SAS6 are surrounded by a SAS4-toroid in the lumen of the microtubule wall. Sequential nucleation of axonemes and mitotic spindles is associated with a dynamic movement of γ-tubulin from the basal bodies to the intranuclear bodies. This dynamic architecture relies on two non-canonical regulators, the calcium-dependent protein kinase 4 and the serine/arginine-protein kinase 1. Altogether, these results provide insights into the molecular organisation of a bipartite MTOC that may reflect a functional transition of a basal body to coordinate axoneme assembly with mitosis. Author summary: Malaria parasites rely on multiple phases of proliferation to build large populations from a small number of founding members, both in the human and the mosquito hosts. Malaria transmission relies on the production of gametes that follows ingestion by a mosquito. In particular, the Plasmodium microgametocyte rapidly forms eight flagellated microgametes through an unusual coupling of mitosis and axoneme formation via a single microtubule organization centre (MTOC). Due to evolutionarily divergence of Plasmodium parasites from well-studied organisms and their relatively small size, little is known about the molecular organisation and the dynamics of this dual MTOC. In this study, we have implemented expansion microscopy (ExM) techniques to image microtubule structures in Plasmodium gametocytes. ExM consists in physically expanding the cell while keeping its cellular structures intact to allow nanoscale imaging of fluorescently labelled structures with conventional microscopes. This reveals the highly dynamic molecular organisation of this bipartite MTOC regulated by two non-canonical kinase regulators as well as the requirement of conserved MTOC components to coordinate mitosis and axoneme assembly. Altogether our results allow to better understand how malaria-causing parasites replicate to form large populations and how they form flagella to colonise the mosquito. [ABSTRACT FROM AUTHOR]

Background: Soybean is a major legume crop with high nutritional and environmental values suitable for sustainable agriculture. Noncoding RNAs (ncRNAs), including microRNAs (miRNAs) and long noncoding RNAs (lncRNAs), are important regulators of gene functions in eukaryotes. However, the interactions between these two types of ncRNAs in the context of plant physiology, especially in response to salinity stress, are poorly understood. Results: Here, we challenged a cultivated soybean accession (C08) and a wild one (W05) with salt treatment and obtained their small RNA transcriptomes at six time points from both root and leaf tissues. In addition to thoroughly analyzing the differentially expressed miRNAs, we also documented the first case of miRNA arm-switching (miR166m), the swapping of dominant miRNA arm expression, in soybean in different tissues. Two arms of miR166m target different genes related to salinity stress (chloroplastic beta-amylase 1 targeted by miR166m-5p and calcium-dependent protein kinase 1 targeted by miR166m-3p), suggesting arm-switching of miR166m play roles in soybean in response to salinity stress. Furthermore, two pairs of miRNA:lncRNA interacting partners (miR166i-5p and lncRNA Gmax_MSTRG.35921.1; and miR394a-3p and lncRNA Gmax_MSTRG.18616.1) were also discovered in reaction to salinity stress. Conclusions: This study demonstrates how ncRNA involves in salinity stress responses in soybean by miRNA arm switching and miRNA:lncRNA interactions. The behaviors of ncRNAs revealed in this study will shed new light on molecular regulatory mechanisms of stress responses in plants, and hence provide potential new strategies for crop improvement. [ABSTRACT FROM AUTHOR]

Ca2+/calmodulin-dependent protein kinase kinases (CaMKKα and β) are regulatory kinases for multiple downstream kinases, including CaMKI, CaMKIV, PKB/Akt, and AMP-activated protein kinase (AMPK) through phosphorylation of each activation-loop Thr residue. In this report, we biochemically characterize the oligomeric structure of CaMKK isoforms through a heterologous expression system using COS-7 cells. Oligomerization of CaMKK isoforms was readily observed by treating CaMKK transfected cells with cell membrane permeable crosslinkers. In addition, His-tagged CaMKKα (His–CaMKKα) pulled down with FLAG-tagged CaMKKα (FLAG–CaMKKα) in transfected cells. The oligomerization of CaMKKα was confirmed by the fact that GST–CaMKKα/His–CaMKKα complex from transiently expressed COS-7 cells extracts was purified to near homogeneity by the sequential chromatography using glutathione-sepharose/Ni-sepharose and was observed in a Ca2+/CaM-independent manner by reciprocal pulldown assay, suggesting the direct interaction between monomeric CaMKKα. Furthermore, the His-CaMKKα kinase-dead mutant (D293A) complexed with FLAG–CaMKKα exhibited significant CaMKK activity, indicating the active CaMKKα multimeric complex. Collectively, these results suggest that CaMKKα can self-associate in the cells, constituting a catalytically active oligomer that might be important for the efficient activation of CaMKK-mediated intracellular signaling. • CaMKK isoforms are capable of forming a multimeric complex in cells. • CaMKKα oligomerization is promoted by self-association, not mediated by scaffold proteins. • Promotion of homomeric oligomerization of CaMKKα is in a Ca2+/calmodulin-independent manner. • CaMKKα oligomer is catalytically active. [ABSTRACT FROM AUTHOR]

There is an important activity of 4-thiazolidinone derivatives on the active site of protein kinases of the parasite Toxoplasma gondii. The calcium-dependent protein kinase CDPK1 is an important object of study, due to its activity in host cell invasion and parasite motility. Twelve structural modifications with alkyl, aromatic and bumpedkinase substitutions at position 3 of the 2-hydrazono-4-thiazolidinone derivative were proposed. A molecular docking and ADME-T properties study was carried out, where derivatives 8A, 2B and 3C presented low binding energy scores (-9.51, -9.19, -10.58) Kcal/mol respectively, a good toxicological profile and no violation of Lipinski's rules being candidates for synthesis for in vivo evaluation. [ABSTRACT FROM AUTHOR]

Appropriate flowering time is critical for the reproductive success of plant species. Emerging evidence indicates that calcium may play an important role in the regulation of flowering time. However, the underlying molecular mechanisms remain unclear. In this study, we demonstrate that calcium-dependent protein kinase 32 (CPK32) regulates flowering time by affecting the alternative polyadenylation of FLOWERING CONTROL LOCUS A (FCA) and altering the transcription of FLOWERING LOCUS C (FLC), a central repressor of flowering time. The knockdown of CPK32 results in an obvious late flowering phenotype and dramatically enhanced FLC transcription. CPK32 interacts with FCA, and phosphorylates the serine592 of FCA in a Ca2+-dependent manner. Moreover, the ratio of abundance of the FCA transcripts (FCA-D and FCA-P) changes significantly in the cpk32 mutant, which subsequently affects FLC expression and consequently regulates floral transition. The present evidence demonstrates that CPK32 modulates flowering time by regulating FCA alternative polyadenylation and consequent FLC expression. [ABSTRACT FROM AUTHOR]

Background: Escherichia coli produces Shiga toxin (Stx), a pentamer composed of one A subunit and four B subunits. The B subunit of Stx (StxB) mediated the attachment of the holotoxin to the cell surface while the A subunit (StxA) has N-glycosidase activity, resulting in protein synthesis and cell death inhibition. Stx-induced cytotoxicity and apoptosis have been observed in various cell lines, although the signaling effectors are not precisely defined. Activated by protein kinases (PK), the signaling pathway in human tumors plays an oncogenic role. Tumor proliferation, survival, and metastasis are promoted by kinase receptors. In this regard, PK regulatory effects on the cellular constituents of the tumor microenvironment can affect immunosuppressive purposes. Methods: In this study, kinase inhibitors were used to evaluate the influence of Stx and its subunits on HeLa and Vero cells. Selective inhibitors of protein kinase C (PKC), CaM kinase (calmodulin kinase), protein kinase A (PKA), and protein kinase G (PKG) were used to compare the signaling activity of each subunit. Results: The ribotoxic activity in the target cells will lead to rapid protein synthesis inhibition and cell death in the mammalian host. The expression of Bcl2 family members was also assessed. Protein kinase signaling by Stx and its A and B subunits was induced by PKA, PKG, and PKC in HeLa cells. CaM kinase induction was significant in Vero cells. StxB significantly induced the pro-apoptotic Bax signaling factor in HeLa cells. Conclusion: The assessment of different signaling pathways utilized by Stx and its subunits could help in a better understanding of various cell death responses. The use of inhibitors can block cell damage and disease progression and create therapeutic compounds for targeted cancer therapy. Inhibition of these pathways is the primary clinical goal. [ABSTRACT FROM AUTHOR]

In this work, 21 dihydroartemisinin derivatives were synthesized, their chemical structures were characterized by 1H NMR, 13H NMR and high-resolution MS techniques, and anti-Toxoplasma gondii activity in vitro was evaluated using thiazole blue (MTT) assay. The selectivity index of compound (3R,5aS,6R,8aS,9R,12R,12aR)-3,6,9-trime-thyldeca-hydro-12H-3,12-epoxy[1,2]dioxepino[4,3-i]isochromen-10-yl 4-oxo-4-(pyridin-4-ylamino)butanoate (E2) was 2.24, which showed the strongest anti-T. gondii activity. In addition, the results of molecular docking studies show that E2 can be a better inhibitor of T. gondii calcium-dependent protein kinase 1 (TgCDPK1). Therefore, compound E2 has good potential as a drug for T. gondii treatment, and further research is needed to clarify its mechanism of action. [ABSTRACT FROM AUTHOR]

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This SR calcium leak can give rise to calcium overload and calcium waves. Subsequent calcium-dependent activation of the sodium/calcium-exchange current (INCX) and calcium-activated chloride current [ICl(Ca)] produces a transient inward current that determines the amplitude of the delayed afterdepolarization (DAD). Finally, there is evidence that diastolic SR calcium release (e.g. promoted by caffeine) can acutely inhibit cardiac sodium channels, which would also be expected to reduce the likelihood of triggered arrhythmias[10] ( I Graphical Abstract i ). [Extracted from the article]

Verticillium dahliae is a soil-borne fungus that causes vascular wilt through the roots of plants. Verticillium wilt caused by V. dahliae is one of the main diseases in cotton producing areas of the world, resulting in huge economic losses. Breeding resistant varieties is the most economical and effective method to control Verticillium wilt. Calcium-dependent protein kinases (CDPKs) play a pivotal role in plant innate immunity, including regulation of oxidative burst, gene expression as well as hormone signal transduction. However, the function of cotton CDPKs in response to V. dahliae stress remains unexplored. In this study, 96, 44 and 57 CDPKs were identified from Gossypium hirsutum , Gossypium raimondii and Gossypium arboretum , respectively. Phylogenetic analysis showed that these CDPKs could be divided into four branches. All GhCDPKs of the same clade are generally similar in gene structure and conserved domain arrangement. Cis-acting elements related to hormones, stress response, cell cycle and development were predicted in the promoter region. The expression of GhCDPKs could be regulated by various stresses. Gh_D11G188500.1 and Gh_A11G186100.1 was up-regulated under Vd0738 and Vd991 stress. Further phosphoproteomics analysis showed that Gh_A11G186100.1 (named as GhCDPK28-6) was phosphorylated under the stress of V. dahliae. Knockdown of GhCDPK28-6 expression, the content of reactive oxygen species was increased, a series of defense responses were enhanced, and the sensitivity of cotton to V. dahliae was reduced. Moreover, overexpression of GhCDPK28-6 in Arabidopsis thaliana weakened the resistance of plants to this pathogen. Subcellular localization revealed that GhCDPK28-6 was localized in the cell membrane. We also found that GhPBL9 and GhRPL12C may interact with GhCDPK28-6. These results indicate that GhCDPK28-6 is a potential molecular target for improving resistance to Verticillium wilt in cotton. This lays a foundation for breeding disease-resistant varieties. [ABSTRACT FROM AUTHOR]

Upon pathogen recognition, a transient rise in cytoplasmic calcium levels is one of the earliest events in plants and a prerequisite for defense initiation and signal propagation from a local site to systemic plant tissues. However, it is unclear if calcium signaling differs in the context of priming: Do plants exposed to a first pathogen stimulus and have consequently established systemic acquired resistance (SAR) display altered calcium responses to a second pathogen stimulus? Several calcium indicator systems including aequorin, YC3.6 or R-GECO1 have been used to document local calcium responses to the bacterial flg22 peptide but systemic calcium imaging within a single plant remains a technical challenge. Here, we report on an experimental approach to monitor flg22-induced calcium responses in systemic leaves of primed plants. The calcium-dependent protein kinase CPK5 is a key calcium sensor and regulator of the NADPH oxidase RBOHD and plays a role in the systemic calcium-ROS signal propagation. We therefore compared flg22-induced cytoplasmic calcium changes in Arabidopsis wild-type, cpk5 mutant and CPK5-overexpressing plants (exhibiting constitutive priming) by introgressing the calcium indicator R-GECO1-mTurquoise that allows internal normalization through mTurquoise fluorescence. Aequorin-based analyses were included for comparison. Based on the R-GECO1-mTurquoise data, CPK5-OE appears to reinforce an "oscillatory-like" Ca2+ signature in flg22-treated local tissues. However, no change was observed in the flg22-induced calcium response in the systemic tissues of plants that had been pre-challenged by a priming stimulus – neither in wild-type nor in cpk5 or CPK5-OE-lines. These data indicate that the mechanistic manifestation of a plant immune memory in distal plant parts required for enhanced pathogen resistance does not include changes in rapid calcium signaling upstream of CPK5 but rather relies on downstream defense responses. [ABSTRACT FROM AUTHOR]

Background: Frogeye leaf spot (FLS) is a destructive fungal disease that affects soybean production. The most economical and effective strategy to control FLS is the use of resistant cultivars. However, the use of a limited number of resistant loci in FLS management will be countered by the emergence of new high-virulence Cercospora sojina races. Therefore, we identified quantitative trait loci (QTL) that control resistance to FLS and identified novel resistant genes using a genome-wide association study (GWAS) on 234 Chinese soybean cultivars. Results: A total of 30,890 single nucleotide polymorphism (SNP) markers were used to estimate linkage disequilibrium (LD) and population structure. The GWAS results showed four loci (p < 0.0001) distributed over chromosomes (Chr.) 5 and 20, that are significantly associated with FLS resistance. No previous studies have reported resistance loci in these regions. Subsequently, 45 genes in the two resistance-related haplotype blocks were annotated. Among them, Glyma20g31630 encoding pyruvate dehydrogenase (PDH), Glyma05g28980, which encodes mitogen-activated protein kinase 7 (MPK7), and Glyma20g31510, Glyma20g31520 encoding calcium-dependent protein kinase 4 (CDPK4) in the haplotype blocks deserves special attention. Conclusions: This study showed that GWAS can be employed as an effective strategy for identifying disease resistance traits in soybean and narrowing SNPs and candidate genes. The prediction of candidate genes in the haplotype blocks identified by disease resistance loci can provide a useful reference to study systemic disease resistance. [ABSTRACT FROM AUTHOR]

Background: Members of the transient receptor potential canonical (TRPC) protein family are widely distributed in the hippocampus of mammals and exert respective and cooperative influences on the functions of neurons. The relationship between specific TRPC subtypes and neuroinflammation is receiving increasing attention.Methods: Using Cx3cr1CreERIL-10-/- transgenic mice and their littermates to study the relationship between TRPC channels and memory impairment.Results: We demonstrated that Cx3cr1CreERIL-10-/- mice displayed spatial memory deficits in object location recognition (OLR) and Morris water maze (MWM) tasks. The decreased levels of TRPC4 and TRPC5 in the hippocampal regions were verified via reverse transcription polymerase chain reaction, western blotting, and immunofluorescence tests. The expression of postsynaptic density protein 95 (PSD95) and synaptophysin in the hippocampus decreased with an imbalance in the local inflammatory environment in the hippocampus. The number of cells positive for ionized calcium-binding adaptor molecule 1 (Iba1), a glial fibrillary acidic protein (GFAP), increased with the high expression of interleukin 6 (IL-6) in Cx3cr1CreERIL-10-/- mice. The nod-like receptor protein 3 (NLRP3) inflammasome was also involved in this process, and the cytokines IL-1β and IL-18 activated by NLRP3 were also elevated by western blotting. The co-localization of TRPC5 and calmodulin-dependent protein kinase IIα (CaMKIIα) significantly decreased TRPC5 expression in excitatory neurons. AAV9-CaMKIIα-TRPC5 was used to upregulate TRPC5 in excitatory neurons in the hippocampus.Conclusions: The results showed that the upregulation of TRPC5 improved the memory performance of Cx3cr1CreERIL-10-/- mice related to inhibiting NLRP3 inflammasome-associated neuroinflammation. [ABSTRACT FROM AUTHOR]

Systemic acquired resistance (SAR) in plants is a defense response that provides resistance against a wide range of pathogens at the whole-plant level following primary infection. Although the molecular mechanisms of SAR have been extensively studied in recent years, the role of phosphorylation that occurs in systemic leaves of SAR-induced plants is poorly understood. We used a data-independent acquisition (DIA) phosphoproteomics platform based on high-resolution mass spectrometry in an Arabidopsis thaliana model to identify phosphoproteins related to SAR establishment. A total of 8011 phosphorylation sites from 3234 proteins were identified in systemic leaves of Pseudomonas syringae pv. maculicola ES4326 (Psm ES4326) and mock locally inoculated plants. A total of 859 significantly changed phosphoproteins from 1119 significantly changed phosphopeptides were detected in systemic leaves of Psm ES4326 locally inoculated plants, including numerous transcription factors and kinases. A variety of defense response-related proteins were found to be differentially phosphorylated in systemic leaves of Psm ES4326 locally inoculated leaves, suggesting that these proteins may be functionally involved in SAR through phosphorylation or dephosphorylation. Significantly changed phosphoproteins were enriched mainly in categories related to response to abscisic acid, regulation of stomatal movement, plant–pathogen interaction, MAPK signaling pathway, purine metabolism, photosynthesis-antenna proteins, and flavonoid biosynthesis. A total of 28 proteins were regulated at both protein and phosphorylation levels during SAR. RT-qPCR analysis revealed that changes in phosphorylation levels of proteins during SAR did not result from changes in transcript abundance. This study provides comprehensive details of key phosphoproteins associated with SAR, which will facilitate further research on the molecular mechanisms of SAR. [ABSTRACT FROM AUTHOR]

Background: The intricate relationship between type 2 diabetes mellitus (T2DM) and Alzheimer's disease (AD) suggests that insulin is involved in modulating AD-related proteins. Alpha-lipoic acid (ALA) can improve insulin resistance (IR) in diabetic rats. However, the role of ALA in alleviating the cognitive decline of T2DM is not yet clear. This study examined the ameliorative effect of ALA on cognitive impairment, cerebral IR, and synaptic plasticity abnormalities in high-fat diet (HFD) plus streptozotocin (STZ) induced diabetic rats. Methods: The HFD/STZ-induced T2DM male Wistar rats were orally administered with ALA (50, 100, or 200 mg/kg BW) once a day for 13 weeks. Abilities of cognition were measured with a passive avoidance test and Morris water maze. Specimens of blood and brain were collected for biochemical analysis after the rats were sacrificed. Western blotting was used to determine protein expressions in the hippocampus and cortex in the insulin signaling pathways, long-term potentiation (LTP), and synaptic plasticity-related protein expressions. Results: Alpha-lipoic acid improved hyperinsulinemia and the higher levels of free fatty acids of the T2DM rats. Behavioral experiments showed that the administration of ALA improved cognitive impairment in HFD/STZ-induced T2DM rats. ALA ameliorated insulin-related pathway proteins [phosphoinositide 3-kinase (PI3K), phospho-protein kinase B (pAkt)/Akt, and insulin-degrading enzyme (IDE)] and the LTP pathway, as well as synaptic plasticity proteins (calmodulin-dependent protein kinase II, cyclic AMP response element-binding protein, and postsynaptic density protein-95) of the cerebral cortex or hippocampus in HFD/STZ-induced T2DM rats. Conclusion: Our findings suggested that ALA may ameliorate cognition impairment via alleviating cerebral IR improvement and cerebral synaptic plasticity in diabetic rats. [ABSTRACT FROM AUTHOR]

Background: Plant annexins are calcium- and lipid-binding proteins that have multiple functions, and a significant amount of research on plant annexins has been reported in recent years. However, the functions of annexins in diverse biological processes in rice are largely unclear. Results: Herein, we report that OsANN4, a calcium-binding rice annexin protein, was induced by abscisic acid (ABA). Under ABA treatment, the plants in which OsANN4 was knocked down by RNA interference showed some visible phenotypic changes compared to the wild type, such as a lower rooting rate and shorter shoot and root lengths. Moreover, the superoxide dismutase (SOD) and catalase (CAT) activities of the RNAi lines were significantly lower and further resulted in higher accumulation of O2.- and H2O2 than those of the wild-type. A Non-invasive Micro-test Technology (NMT) assay showed that ABA-induced net Ca2+ influx was inhibited in OsANN4 knockdown plants. Interestingly, the phenotypic differences caused by ABA were eliminated in the presence of LaCl3 (Ca2+ channel inhibitor). Apart from this, we demonstrated that OsCDPK24 interacted with and phosphorylated OsANN4. When the phosphorylated serine residue of OsANN4 was substituted by alanine, the interaction between OsANN4 and OsCDPK24 was still observed, however, both the conformation of OsANN4 and its binding activity with Ca2+ might be changed. Conclusions: OsANN4 plays a crucial role in the ABA response, partially by modulating ROS production, mediating Ca2+ influx or interacting with OsCDPK24. [ABSTRACT FROM AUTHOR]

CONNECTION PROTEINS TO SYNAPSES AND CIRCUITS These proteins are all integrated into synapses, which are the building blocks of neuronal circuits. Initially, most studies of the mechanisms of memory focused on excitatory synapses, but recently it has become clear that we must also consider the plasticity of inhibitory GABAergic synapses in memory and circuit excitability. IDENTIFYING THE PLAYERS: THE PROTEINS OF MEMORY Several synaptic proteins lead to memory impairment when knocked-out or inhibited suggesting that they have a role in the molecular mechanisms of memory. The nature of memory has been a source of fascination since at least the ancient Greek philosophers and likely longer. [Extracted from the article]

Abstract Background Drought is one of the most adverse environmental factors limiting crop productions and it is important to identify key genetic determinants for food safety. Calcium-dependent protein kinases (CPKs) are known to be involved in plant growth, development, and environmental stresses. However, biological functions and regulatory mechanisms of many plant CPKs have not been explored. In our previous study, abundance of the wheat CPK34 (TaCPK34) protein was remarkably upregulated in wheat plants suffering from drought stress, inferring that it could be involved in this stress. Therefore, here we further detected its function and mechanism in response to drought stress. Results Transcripts of the TaCPK34 gene were significantly induced after PEG-stimulated water deficiency (20% PEG6000) or 100 μM abscisic acid (ABA) treatments. The TaCPK34 gene was transiently silenced in wheat genome by using barley stripe mosaic virus-induced silencing (BSMV-VIGS) method. After 14 days of drought stress, the transiently TaCPK34-silenced wheat seedlings showed more sensitivity compared with control, and the plant biomasses and relative water contents significantly decreased, whereas soluble sugar and MDA contents increased. The iTRAQ-based quantitative proteomics was employed to measure the protein expression profiles in leaves of the transiently TaCPK34-silenced wheat plants after drought stress. There were 6103 proteins identified, of these, 51 proteins exhibited significantly altered abundance, they were involved in diverse function. And sequence analysis on the promoters of genes, which encoded the above identified proteins, indicated that some promoters harbored some ABA-responsive elements. We determined the interactions between TaCPK34 and three identified proteins by using bimolecular fluorescent complementation (BiFC) method and our data indicated that TaCPK34directly interacted with the glutathione S-transferase 1 and prx113, respectively. Conclusions Our study suggested that the TaCPK34 gene played positive roles in wheat response to drought stress through directly or indirectly regulating the expression of ABA-dependent manner genes, which were encoding identified proteins from iTRAQ-based quantitative proteomics. And it could be used as one potential gene to develop crop cultivars with improved drought tolerance.

Verticillium dahliae is a soil-borne fungus that causes vascular wilt through the roots of plants. Verticillium wilt caused by V. dahliae is one of the main diseases in cotton producing areas of the world, resulting in huge economic losses. Breeding resistant varieties is the most economical and effective method to control Verticillium wilt. Calcium-dependent protein kinases (CDPKs) play a pivotal role in plant innate immunity, including regulation of oxidative burst, gene expression as well as hormone signal transduction. However, the function of cotton CDPKs in response to V. dahliae stress remains unexplored. In this study, 96, 44 and 57 CDPKs were identified from Gossypium hirsutum, Gossypium raimondii and Gossypium arboretum, respectively. Phylogenetic analysis showed that these CDPKs could be divided into four branches. All GhCDPKs of the same clade are generally similar in gene structure and conserved domain arrangement. Cis-acting elements related to hormones, stress response, cell cycle and development were predicted in the promoter region. The expression of GhCDPKs could be regulated by various stresses. Gh_D11G188500.1 and Gh_A11G186100.1 was up-regulated under Vd0738 and Vd991 stress. Further phosphoproteomics analysis showed that Gh_A11G186100.1 (named as GhCDPK28-6) was phosphorylated under the stress of V. dahliae. Knockdown of GhCDPK28-6 expression, the content of reactive oxygen species was increased, a series of defense responses were enhanced, and the sensitivity of cotton to V. dahliae was reduced. Moreover, overexpression of GhCDPK28-6 in Arabidopsis thaliana weakened the resistance of plants to this pathogen. Subcellular localization revealed that GhCDPK28-6 was localized in the cell membrane. We also found that GhPBL9 and GhRPL12C may interact with GhCDPK28-6. These results indicate that GhCDPK28-6 is a potential molecular target for improving resistance to Verticillium wilt in cotton. This lays a foundation for breeding disease-resistant varieties.

Calcium-dependent protein kinases (CDPKs) regulate plant development and stress responses. However, the interaction of these protein kinases with the abscisic acid (ABA) stress hormone signalling system has not been studied in detail. In Arabidopsis, AtCPK1 plays an important role in the acclimation of plants to environmental stresses. Phylogenetic and molecular analyses showed that, among 50 isoforms of Glycine max (L.) Merrill CDPKs, the GmCDPK27/GmCDPK48, GmCDPK5/GmCDPK24, and GmCDPK10/GmCDPK46 paralogous pairs were the isoforms most related to AtCDPK1. We investigated the expression of the corresponding six GmCDPKs genes during treatment with cold, heat, and salt stress. Wild soybean was the most resistant to stresses, and among the three cultivars studied (Sfera, Hodgson, and Hefeng25), Sfera was close to the wild type in terms of resistance. GmCDPK5 and GmCDPK10 were the most responsive to stress treatments, especially in wild soybean, compared with cultivars. Among the studied GmCDPK isoforms, only GmCDPK5 expression increased after treatment with abscisic acid (ABA) in a dose- and time-dependent manner. Targeted LC-MS/MS analysis of endogenous ABA levels showed that wild soybean and Sfera had nearly twice the ABA content of Hodgson and Hefeng25. An analysis of the expression of marker genes involved in ABA biosynthesis showed that GmNCED1-gene-encoding 9-cis-epoxycarotenoid dioxygenase 1 is induced to the greatest extent in wild soybean and Sfera under salt, cold, and heat exposure. Our data established a correlation between the induction of GmCDPK5 and ABA biosynthesis genes. GmCDPK5 is an interesting target for genetic and bioengineering purposes and can be used for genetic editing, overexpression, or as a marker gene in soybean varieties growing under unfavourable conditions.

Cell signaling pathways regulating myosin regulatory light chain (LC20) phosphorylation contribute to determining contractile responses in smooth muscles. Following excitation and contraction, phasic smooth muscles, such as the digestive tract and urinary bladder, undergo relaxation due to a decline of cellular Ca²þ concentration and decreased Ca²þ sensitivity of LC20 phosphorylation, named Ca²þ desensitization. Here, we determined the mechanisms underlying the temporal Ca²þ desensitization of LC20 phosphorylation in phasic smooth muscles using permeabilized strips of the mouse ileum and urinary bladder. Upon stimulation with pCa6.0 at 20°C, contraction and LC20 phosphorylation peaked within 30 s and then declined to about 50% of the peak force at 2min after stimulation. During the relaxation phase after the contraction, LC20 kinase [myosin light chain kinase (MLCK)] was inactivated, but no fluctuation in LC20 phosphatase activity occurred, suggesting that MLCK inactivation is a cause of the Ca²þ-induced Ca²þ desensitization of LC20 phosphorylation. MLCK inactivation was associated with phosphorylation at the calmodulin-binding domain of the kinase. Treatment with STO-609 and TIM-063 antagonists for Ca²þ/calmodulin (CaM)-dependent protein kinase kinase-b (CaMKKb) attenuated both the phasic response of the contraction and MLCK phosphorylation, whereas neither CaM kinase II, AMP-activated protein kinase, nor p21-activated kinase induced MLCK inactivation in phasic smooth muscles. Conversely, protein phosphatase 2A inhibition amplified the phasic response. Signaling pathways through CaMKKb and protein phosphatase 2A may contribute to regulating the phasic response of smooth muscle contraction. [ABSTRACT FROM AUTHOR]

Protein acetylation is a reversible posttranslational modification, which is regulated by lysine acetyltransferase (KAT) and lysine deacetyltransferase (KDAC). Although protein acetylation has been shown to regulate synaptic plasticity, this was mainly for histone protein acetylation. The function and regulation of nonhistone protein acetylation in synaptic plasticity and learning remain largely unknown. Calmodulin (CaM), a ubiquitous Ca2+ sensor, plays critical roles in synaptic plasticity such as long-term potentiation (LTP). During LTP induction, activation of NMDA receptor triggers Ca2+ influx, and the Ca2+ binds with CaM and activates calcium/ calmodulin-dependent protein kinase IIα (CaMKIIα). In our previous study, we demonstrated that acetylation of CaM was important for synaptic plasticity and fear learning in mice. However, the KAT responsible for CaM acetylation is currently unknown. Here, following an HEK293 cell-based screen of candidate KATs, steroid receptor coactivator 3 (SRC3) is identified as the most active KAT for CaM. We further demonstrate that SRC3 interacts with and acetylates CaM in a Ca2+ and NMDA receptor-dependent manner. We also show that pharmacological inhibition or genetic downregulation of SRC3 impairs CaM acetylation, synaptic plasticity, and contextual fear learning in mice. Moreover, the effects of SRC3 inhibition on synaptic plasticity and fear learning could be rescued by 3KQ-CaM, a mutant form of CaM, which mimics acetylation. Together, these observations demonstrate that SRC3 acetylates CaM and regulates synaptic plasticity and learning in mice. [ABSTRACT FROM AUTHOR]

Long‐term potentiation (LTP) is a highly studied cellular process, yet determining the transduction and gamma aminobutyric acid (GABAergic) pathways that are the essential versus modulatory for LTP elicited by theta burst stimulation (TBS) in the hippocampal Cornu Ammonis 1 (CA1) area is still elusive, due to the use of different TBS intensities, patterns or different rodent/cellular models. We now characterised the developmental maturation and the transduction and GABAergic pathways required for mild TBS‐induced LTP in hippocampal CA1 area in male rats. LTP induced by TBS (5x4) (five bursts of four pulses delivered at 100 Hz) lasted for up to 3 h and was increasingly larger from weaning to adulthood. Stronger TBS patterns ‐ TBS (15x4) or three TBS (15x4) separated by 6 min induced nearly maximal LTP not being the best choice to study the value of LTP‐enhancing drugs. LTP induced by TBS (5x4) in young adults was fully dependent on N‐methyl D‐aspartate (NMDA) receptor and calmodulin‐dependent protein kinase II (CaMKII) activity but independent of protein kinase A (PKA) or protein kinase C (PKC) activity. Furthermore, it was partially dependent on GABAB receptor activation and was potentiated by GABAA receptor blockade and less by GAT‐1 transporter blockade. AMPA GluA1 phosphorylation on Ser831 (CaMKII target) but not GluA1 Ser845 (PKA target) was essential for LTP expression. The phosphorylation of the Kv4.2 channel was observed at Ser438 (CaMKII target) but not at Thr602 or Thr607 (ERK/MAPK pathway target). This suggests that cellular kinases like PKA, PKC, or kinases of the ERK/MAPK family although important modulators of TBS (5x4)‐induced LTP may not be essential for its expression in the CA1 area of the hippocampus. [ABSTRACT FROM AUTHOR]

Background: Babesia bovis is one of the most significant tick-transmitted pathogens of cattle worldwide. Babesia bovis parasites have a complex lifecycle, including development within the mammalian host and tick vector. Each life stage has developmental forms that differ in morphology and metabolism. Differentiation between these forms is highly regulated in response to changes in the parasite's environment. Understanding the mechanisms by which Babesia parasites respond to environmental changes and the transmission cycle through the biological vector is critically important for developing bovine babesiosis control strategies. Results: In this study, we induced B. bovis sexual stages in vitro using xanthurenic acid and documented changes in morphology and gene expression. In vitro induced B. bovis sexual stages displayed distinctive protrusive structures and surface ruffles. We also demonstrated the upregulation of B. bovis calcium-dependent protein kinase 4 (cdpk4), tubulin-tyrosine ligase (ttl), and methyltransferase (mt) genes by in vitro induced sexual stages and during parasite development within tick midguts. Conclusions: Similar to other apicomplexan parasites, it is likely that B. bovis upregulated genes play a vital role in sexual reproduction and parasite transmission. Herein, we document the upregulation of cdpk4, ttl, and mt genes by both B. bovis in vitro induced sexual stages and parasites developing in the tick vector. Understanding the parasite's biology and identifying target genes essential for sexual reproduction will enable the production of non-transmissible live vaccines to control bovine babesiosis. [ABSTRACT FROM AUTHOR]

The ability of an organism to sense and respond to environmental redox fluctuations relies on a signaling network that is incompletely understood in apicomplexan parasites such as Toxoplasma gondii. The impact of changes in redox upon the development of this intracellular parasite is not known. Here, we provide a revised collection of 58 genes containing domains related to canonical antioxidant function, with their encoded proteins widely dispersed throughout different cellular compartments. We demonstrate that addition of exogenous H2O2 to human fibroblasts infected with T. gondii triggers a Ca2+ flux in the cytosol of intracellular parasites that can induce egress. In line with existing models, egress triggered by exogenous H2O2 is reliant upon both Calcium-Dependent Protein Kinase 3 and diacylglycerol kinases. Finally, we show that the overexpression a glutaredoxin-roGFP2 redox sensor fusion protein in the parasitophorous vacuole severely impacts parasite replication. These data highlight the rich redox network that exists in T. gondii , evidencing a link between extracellular redox and intracellular Ca2+ signaling that can culminate in parasite egress. Our findings also indicate that the redox potential of the intracellular environment contributes to normal parasite growth. Combined, our findings highlight the important role of redox as an unexplored regulator of parasite biology. [ABSTRACT FROM AUTHOR]

Drought is a major factor which reduces cane growth and productivity. In the present study, we sequenced drought susceptible (V1) and drought tolerant (V2) sugarcane varieties using high-throughput miRNA deep sequencing method to study the regulation of gene expression by miRNAs during drought stress in sugarcane. A total of 1224 conserved miRNAs which belong to 89 miRNA families were identified and 38% of the differentially regulated miRNAs were common for both varieties. Additionally 435 novel miRNAs were also identified from four small RNA libraries. We identified 145 miRNAs that were differentially expressed in susceptible variety (V1–31) and 143 miRNAs differentially expressed in the tolerant variety (V2–31). Target prediction revealed that the genes mainly encoded transcription factors, proteins, phosphatase and kinases involved in signal transduction pathways, integral component of membrane and inorganic ion transport metabolism, enzymes involved in carbohydrate transport and metabolism and drought-stress-related proteins involved in defense mechanisms. Pathway analysis of targets revealed that "General function prediction only" was the most significant pathway observed in both tolerant and susceptible genotypes followed by "signal transduction mechanisms". Functional annotation of the transcripts revealed genes like calcium-dependent protein kinase, respiratory burst oxidase, caffeic acid 3-O-methyltransferase, peroxidase, calmodulin, glutathione S-transferase and transcription factors like MYB, WRKY that are involved in drought tolerant pathways. qRT-PCR was used to verify the expression levels of miRNAs and their potential targets obtained from RNA sequencing results. [ABSTRACT FROM AUTHOR]

Apicomplexan parasites, including Toxoplasma gondii (T. gondii), express different types of calcium-dependent protein kinases (CDPKs), which perform a variety of functions, including attacking and exiting the host cells. In the current bioinformatics study, we have used several web servers to predict the basic features and specifications of the CDPK7 protein. The findings showed that CDPK7 protein has 2133 amino acid residues with an average molecular weight (MW) of 219085.79 D. The aliphatic index with 68.78 and grand average of hydropathicity (GRAVY) with -0.331 score were estimated. The outcomes of current research showed that the CDPK7 protein included 502 alpha-helix, 1311 random coils, and 320 extended strands with GOR4 method. Considering the Ramachandran plot, the favored region contains more than 92% of the amino acid residues. In addition, evaluation of antigenicity and allergenicity showed that CDPK7 protein has immunogenic and nonallergenic nature. The present research provides key data for more animal-model study on the CDPK7 protein to design an efficient vaccine against toxoplasmosis in the future. [ABSTRACT FROM AUTHOR]

Purpose: To identify some natural molecules (inhibitors) against avian coccidia (Eimeria tenella) through molecular docking in order to find the candidate drugs for poultry industry. Methods: The natural compounds, 6’-de-O-acetylcupacinoside, apigenin, artemisinin, cupacinoside, quercetin and rutin, were screened on the basis of previous research data. These biomolecules were selected as potent inhibitors based on extant literature on calcium-dependent protein kinases (CDPKs) in Eimeria tenella. Results: All the compounds showed good interactions with 6’-de-O-acetylcupacinoside. Based on docking studies, quercetin produced the best interaction, with a binding energy of -7.04 kcal/mol, when compared to the other compounds. Conclusions: Based on these in silico results, the screened compounds have great potential for use against E. tenella. In particular, the results suggest that quercetin may be beneficial in the control of avian coccidia through its strong inhibitory potential on CDPKs of E. tenella. [ABSTRACT FROM AUTHOR]

Background: Post-stroke depression (PSD) is one of the most common complications of stroke. Many studies have confirmed that PSD is associated with the abnormal release of glutamic acid (Glu) via the Ca2+/calmodulin (CaM)/CaM-dependent protein kinase II (CaMKII) and connexin 43 (Cx43) hemichannel pathways. Onjisaponin B, a traditional Chinese medicine, has been discovered to have neuroprotective effects. Objectives: In this study, we aimed to investigate the effects and underlying mechanisms of onjisaponin B on the release of glutamate in rat astrocytes subjected to oxygen and glucose deprivation (OGD). Materials and Methods: The ischemic and anoxic cell model was established in rat astrocytes through OGD injury. Rat astrocytes were randomly divided into control, model, 10 μM onjisaponin B, and 20 μM onjisaponin B groups. Cell viability was assessed by acridine orange/ethidium bromide bilabel assay. The intracellular Ca2+ level was measured with Fluo-3-AM as the fluorescence indicator. Western blot analysis and quantitative polymerase chain reaction were used to detect the expressions of Cx43 and CaMKII in the samples. The glutamate level of the extracellular fluid was determined using high-performance liquid chromatography-mass spectrometry. Results: The intracellular Ca2+ levels and the mRNA and protein expression of CaMKII and Cx43 in the onjisaponin B group were significantly lower than that of the model group. The glutamate level in the extracellular fluid was significantly reduced by onjisaponin B in comparison to that in the model group. Moreover, onjisaponin B attenuated the inhibitory effect of OGD on the cell viability of astrocytes. Conclusion: The results of this study suggest that onjisaponin B reduced the release of glutamate via inhibition of the Ca2+/CaMKII and Cx43 pathways in the rat astrocytes, thus inhibiting the downstream glutamic pathway and reinforcing the cell viability of astrocyte. [ABSTRACT FROM AUTHOR]

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As essential calcium ion (Ca2+) sensors in plants, calcium-dependent protein kinases (CDPKs) function in regulating the environmental adaptation of plants. However, the response mechanism of CDPKs to salt stress is not well understood. In the current study, the wheat salt-responsive gene TaCDPK27 was identified. The open reading frame (ORF) of TaCDPK27 was 1875 bp, coding 624 amino acids. The predicted molecular weight and isoelectric point were 68.905 kDa and 5.6, respectively. TaCDPK27 has the closest relationship with subgroup III members of the CDPK family of rice. Increased expression of TaCDPK27 in wheat seedling roots and leaves was triggered by 150 mM NaCl treatment. TaCDPK27 was mainly located in the cytoplasm. After NaCl treatment, some of this protein was transferred to the membrane. The inhibitory effect of TaCDPK27 silencing on the growth of wheat seedlings was slight. After exposure to 150 mM NaCl for 6 days, the NaCl stress tolerance of TaCDPK27-silenced wheat seedlings was reduced, with shorter lengths of both roots and leaves compared with those of the control seedlings. Moreover, silencing of TaCDPK27 further promoted the generation of reactive oxygen species (ROS); reduced the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT); aggravated the injury to photosystem II (PS II); and increased programmed cell death (PCD) in wheat leaves under NaCl treatment, confirming that the TaCDPK27-silenced seedlings exhibited more NaCl injury than control seedlings. Taken together, the decrease in NaCl tolerance in TaCDPK27-silenced seedlings was due to excessive ROS accumulation and subsequent aggravation of the NaCl-induced PCD. TaCDPK27 may be essential for positively regulating salt tolerance in wheat seedlings.

This study aimed to evaluate the acute effects of 2.856 GHz and 1.5 GHz microwaves on spatial memory and cAMP response element binding (CREB)-related pathways. A total of 120 male Wistar rats were divided into four groups: a control group (C); 2.856 GHz microwave exposure group (S group); 1.5 GHz microwave exposure group (L group); and 2.856 and 1.5 GHz cumulative exposure group (SL group). Decreases in spatial memory abilities, changes in EEG, structural injuries, and the downregulation of phosphorylated-Ak strain transforming (p-AKT), phosphorylated-calcium/calmodulin-dependent protein kinase II (p-CaMKII), phosphorylated extracellular signal regulated kinase (p-ERK) and p-CREB was observed 6 h after microwave exposure. Significant differences in the expression of p-CaMKII were found between the S and L groups. The power amplitudes of the EEG waves (θ, δ), levels of structural injuries and the expression of p-AKT, p-CaMK II, p-CREB, and p-ERK1/2 were significantly different in the S and L groups compared to the SL group. Interaction effects between the 2.856 and 1.5 GHz microwaves were found in the EEG and p-CREB changes. Our findings indicated that 2.856 GHz and 1.5 GHz microwave exposure induced a decline in spatial memory, which might be related to p-AKT, p-CaMK II, p-CREB and p-ERK1/2. [ABSTRACT FROM AUTHOR]

It has shown that schizophrenia (SCZ) is associated with a higher chance of myocardial infarction (MI) and increased mortality. However, the underlying mechanism is largely unknown. Here, we first constructed a literature-based genetic pathway linking SCZ and MI, and then we tested the expression levels of the genes involved in the pathway by a meta-analysis using nine gene expression datasets of MI. In addition, a literature-based data mining process was conducted to explore the connection between SCZ at different levels: small molecules, complex molecules, and functional classes. The genetic pathway revealed nine genes connecting SCZ and MI. Specifically, SCZ activates two promoters of MI (IL6 and CRP) and deactivates seven inhibitors of MI (ADIPOQ, SOD2, TXN, NGF, ADORA1, NOS1, and CTNNB1), suggesting that no protective role of SCZ in MI was detected. Meta-analysis showed that one promoter of MI (CRP) presented no significant increase, and six out of seven genetic inhibitors of MI demonstrated minor to moderately increased expression. Therefore, the elevation of CRP and inhibition of the six inhibitors of MI by SCZ could be critical pathways to promote MI. Nine other regulators of MI were influenced by SCZ, including two gene families (inflammatory cytokine and IL1 family), five small molecules (lipid peroxide, superoxide, ATP, ascorbic acid, melatonin, arachidonic acid), and two complexes (CaM kinase 2 and IL23). Our results suggested that SCZ promotes the development and progression of MI at different levels, including genes, small molecules, complex molecules, and functional classes. [ABSTRACT FROM AUTHOR]

The tuberization phenomenon in potatoes (Solanum tuberosum L.) is a highly synchronized morphophysiological process occurring on the underground stolons under the influence of various intrinsic and extrinsic factors. This involves the participation of the phytochrome sensory system, transport of sucrose from source to stolon, and several regulatory pathways including tuber-inducing hormone biosynthetic lipoxygenase (StLOX1) gene and major calcium-mediated signaling pathway genes (StCDPK and StCaM1). This study was aimed to explore the impacts of distinctive qualities of the light-emitting diode (LED) light with a specific wavelength on in vitro tuberization of potato. Single nodal segments of potato 'Kufri Jyoti' were incubated in vitro on the growth medium at a temperature of 22 ± 2 °C and exposed to various combinations of red (R) and blue (B) LED light. The results showed that the combination of 30% red + 70% blue LED light (R30B70) significantly shortened the tuber induction period, increased the number of tubers, and their yield compared with these parameters in the tissues exposed to the white light (W100). The induction of the in vitro tuberization correlated with the enhanced expression of the major tuberizing pathway genes, including the lipoxygenase (StLOX1), calmodulin (CaM1), and calcium-dependent protein kinase (StCDPK). The current study indicated that the combination of red and blue LED lights at R30B70 is the best spectrum for effective in vitro tuberization of potatoes. [ABSTRACT FROM AUTHOR]

Calcium-dependent protein kinases (CDPKs) play essential roles in plant development and stress responses. CDPKs have a conserved kinase domain, followed by an auto-inhibitory junction connected to the calmodulin-like domain that binds Ca2+. These structural features allow CDPKs to decode the dynamic changes in cytoplasmic Ca2+ concentrations triggered by hormones and by biotic and abiotic stresses. In response to these signals, CDPKs phosphorylate downstream protein targets to regulate growth and stress responses according to the environmental and developmental circumstances. The latest advances in our understanding of the metabolic, transcriptional, and protein–protein interaction networks involving CDPKs suggest that they have a direct influence on plant carbon/nitrogen (C/N) balance. In this review, we discuss how CDPKs could be key signaling nodes connecting stress responses with metabolic homeostasis, and acting together with the sugar and nutrient signaling hubs SnRK1, HXK1, and TOR to improve plant fitness. [ABSTRACT FROM AUTHOR]

Calcium (Ca2+)-dependent protein kinases (CDPKs or CPKs) are a unique family of Ca2+ sensor/kinase-effector proteins with diverse functions in plants. In Arabidopsis thaliana, CPK28 contributes to immune homeostasis by promoting degradation of the key immune signaling receptor-like cytoplasmic kinase BOTRYTIS-INDUCED KINASE 1 (BIK1) and additionally functions in vegetative-to-reproductive stage transition. How CPK28 controls these seemingly disparate pathways is unknown. Here, we identify a single phosphorylation site in the kinase domain of CPK28 (Ser318) that is differentially required for its function in immune homeostasis and stem elongation. We show that CPK28 undergoes intermolecular autophosphorylation on Ser318 and can additionally be transphosphorylated on this residue by BIK1. Analysis of several other phosphorylation sites demonstrates that Ser318 phosphorylation is uniquely required to prime CPK28 for Ca2+ activation at physiological concentrations of Ca2+, possibly through stabilization of the Ca2+-bound active state as indicated by intrinsic fluorescence experiments. Together, our data indicate that phosphorylation of Ser318 is required for the activation of CPK28 at low intracellular [Ca2+] to prevent initiation of an immune response in the absence of infection. By comparison, phosphorylation of Ser318 is not required for stem elongation, indicating pathway-specific requirements for phosphorylation-based Ca2+-sensitivity priming. We additionally provide evidence for a conserved function for Ser318 phosphorylation in related group IV CDPKs, which holds promise for biotechnological applications by generating CDPK alleles that enhance resistance to microbial pathogens without consequences to yield. [ABSTRACT FROM AUTHOR]

Objective: Explore the effect of apolipoprotein APOE4 on the hyperphosphorylation of tau protein in mouse hippocampal neurons. Methods: 6-month-old human apolipoprotein APOE3, APOE4 transgenic homozygous mice was used to detect the phosphorylation of tau protein and Calpain protein, CDK5, p35/25 in the hippocampus by western Blot. Ca2+ chelating agent EGTA or dimethyl sulfoxide DMSO was stereotactically injected into the lateral ventricle of the mouse two times with four hours' interval. The mice were sacrificed within two hours after the second administration to detect the changes in the phosphorylation of Calpain protein, CDK5, p35/25 and tau protein in the hippocampus. Results:① Compared to wild-type and APOE3-TR mice, The phosphorylation of tau protein at Ser396, Thr181 and Thr231 sites in the hippocampus of APOE4-TR mice were significantly increased. Meanwhile, the expressions of Calpain 2, p35/25 and CDK5 were also increased. nd. ② Compared with the control group, Ca2+ chelating agent EGTA decreased the phosphorylation of tau protein at Ser396 site in the hippocampus significantly. However, no significant changes were detected in Thr181 and Thr231 sites of phosphorylation of tau protein. Moreover, the expression levels of Calpain 2, p35/25 and CDK5 were also decreased. Conclusion: Human apolipoprotein APOE4 caused abnormally increased phosphorylation of tau protein in the hippocampus of mice. Moreover, the phosphorylation of tau at Ser396 may be regulated by calpain/p35-p25/CDK5 signaling pathway. [ABSTRACT FROM AUTHOR]

Four members of the potato (Solanum tuberosum L.) calcium-dependent protein kinase (CDPK) family StCDPK22/23/24 and StCDPK27, present three functional EF-hands motifs in their calmodulin-like domain (CLD). StCDPK22/23/24 are clustered in clade III-b1 with tomato and Arabidopsis CDPKs that lack the first EF-hand motif, while StCDPK27 is clustered in clade III-b3 with CDPKs that lack EF-hand 2. Members of each clade share similar intron-exon structures and acylation profiles. 3D model predictions suggested that StCDPK22 and StCDPK24 are active kinases that undergo a conformational switch in the presence of Ca2+ even when lacking one functional EF-hand motif; however, assays performed with recombinant proteins indicated that StCDPK24:6xHis was active in all the conditions tested, and its activity was enhanced in the presence of Ca2+, but StCDPK22:6xHis had scarce or null activity. Both kinases share with AtCPK8 the same autophosphorylation pattern in the autoinhibitory (AD) and C-terminal variable (CTV) domains, suggesting that it could be a characteristic of clade III-b1. RT-qPCR analysis revealed that StCDPK22 is mainly expressed in early stages of tuberization, but not limited to, while StCDPK24 expression is more ubiquitous. In silico analysis predicted several abiotic stress-responsive elements in its promoters. Accordingly, StCDPK24 expression peaked at 10 h in in vitro plants exposed to salt shock and then declined. Moreover, a significant increase was observed at 2 h in stems of salt-treated greenhouse plants, suggesting that this CDPK could participate in the early events of the signaling cascade triggered in response to salt. • Calcium-dependent protein kinases are stress sensor-transducers. • StCDPK22 and StCDPK24 present three functional EF-hands in their regulatory domain. • StCDPK22/24 and AtCPK8 share autophosphorylation sites in the autoinhibitory domain. • StCDPK24 could participate in the early events triggered upon salt stress. [ABSTRACT FROM AUTHOR]

Safranal is a free radical scavenger and useful as an antioxidant molecule; however, its promotive role in soybean is not explored. Salt stress decreased soybean growth and safranal improved it even if under salt stress. To study the positive mechanism of safranal on soybean growth, a proteomic approach was used. According to functional categorization, oppositely changed proteins were further confirmed using biochemical techniques. Actin and calcium-dependent protein kinase decreased in soybean root and hypocotyl, respectively, under salt stress and increased with safranal application. Xyloglucan endotransglucosylase/ hydrolase increased in soybean root under salt stress but decreased with safranal application. Peroxidase increased under salt stress and further enhanced by safranal application in soybean root. Actin, RuvB-like helicase, and protein kinase domain-containing protein were upregulated under salt stress and further enhanced by safranal application under salt stress. Dynamin GTPase was downregulated under salt stress but recovered with safranal application under salt stress. Glutathione peroxidase and PfkB domain-containing protein were upregulated by safranal application under salt stress in soybean root. These results suggest that safranal improves soybean growth through the regulation of cell wall and nuclear proteins along with reactive‑oxygen species scavenging system. Furthermore, it might promote salt-stress tolerance through the regulation of membrane proteins involved in endocytosis and post-Golgi trafficking. To study the positive mechanism of safranal on soybean growth, a proteomic approach was used. According to functional categorization, oppositely changed proteins were further confirmed using biochemical techniques. Actin and calcium-dependent protein kinase decreased in soybean root and hypocotyl, respectively, under salt stress and increased with safranal application. Xyloglucan endotransglucosylase/ hydrolase increased in soybean root under salt stress but decreased with safranal application. Peroxidase increased under salt stress and further enhanced by safranal application in soybean root. Actin, RuvB-like helicase, and protein kinase domain-containing protein were upregulated under salt stress and further enhanced by safranal application under salt stress. Dynamin GTPase was downregulated under salt stress but recovered with safranal application under salt stress. Glutathione peroxidase and PfkB domain-containing protein were upregulated by safranal application under salt stress in soybean root. These results suggest that safranal improves soybean growth through the regulation of cell wall and nuclear proteins along with reactive‑oxygen species scavenging system. Furthermore, it might promote salt-stress tolerance through the regulation of membrane proteins involved in endocytosis and post-Golgi trafficking. [Display omitted] • A proteomic approach was used to study the positive mechanism of safranal on soybean growth. • Safranal improves soybean growth through the regulation of cell wall and nuclear proteins. • Safranal regulates reactive‑oxygen species scavenging system. • Safranal promotes salt-stress tolerance through the regulation of membrane proteins. [ABSTRACT FROM AUTHOR]

• Antibiotic-treated (ABX) mice showed impaired memory acquisition in a passive avoidance test. • ABX mice showed decreased hippocampal levels of glutaminase and synaptophysin. • ABX mice showed decreased levels of phosphorylated GluN1, CAMKII, GluA1, and CREB in the hippocampus. • ABX mice showed decreased BDNF, DCX, and NeuN expression levels in the hippocampus. The gut microbiota is associated with memory; however, the relationship between dysbiosis-induced memory deficits and hippocampal glutamatergic neurons remains unclear. In our study, a mouse dysbiosis model showed impaired memory-related behavior in the passive avoidance test; decreased expression levels of glutaminase, excitatory amino acid transporter (EAAT)1, EAAT2, vesicular glutamate transporter 2, synaptophysin, brain-derived neurotrophic factor, doublecortin, neuronal nuclear protein, glial fibrillary acidic protein, and S100β; and decreased phosphorylation of N-methyl-D-aspartate receptor subunit 1, calmodulin-dependent protein kinase II, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunit 1, and cAMP response element-binding protein in the hippocampus. This suggests that dysbiosis-induced memory dysfunction is associated with the hippocampal glutamatergic nervous system. [ABSTRACT FROM AUTHOR]