Calmodulin-dependent Kinase Research Articles

Carvedilol suppresses coronary artery spasm in A-kinase anchoring protein 150 knockout mouse

Abstract Backgrounds A-kinase anchoring proteins (AKAPs) act as scaffold to regulate activation of protein kinases and subsequent downstream signaling. AKAP 150 interacts with protein kinase A and protein kinase C, and has a critical role in regulating L-type calcium channel activity. We previously reported enhanced phospholipase C activity as an important mechanism of coronary spastic angina (CSA), known as Prinzmetal’s angina. Coronary artery spasm was induced by intravenous ergometrine injection in mouse model of vascular smooth muscle cell (VSMC)-specific enhanced phospholipase C activity concomitantly with enhanced intracellular Ca2+ concentration ([Ca2+]i). Then, we hypothesized that coronary artery spasm would be inhibited by AKAP 150 knockout, however, coronary artery spasm was unexpectedly induced in AKAP 150 knockout (AKAP-KO) mice without affecting [Ca2+]i, suggesting that AKAP-KO mouse is an unique model of CSA independent of [Ca2+]i. Previous report showed that AKAP-KO mice have a high Ca2+/calmodulin-dependent protein kinase (CaMK) II activity, and carvedilol, a non-selective beta-adrenergic receptor antagonist, inhibits CaMK II activity. Aim We tested the hypothesis that carvedilol could prevent coronary artery spasm via inhibiting CaMK II activity, which is a key molecule to regulate VSMC contraction, in AKAP-KO mice. Methods and Results We pretreated 14-18-weeks-old AKAP-KO mice with intraperitoneal injection of carvedilol (2.5mg/kg), propranolol (20mg/kg), or vehicle (control). Coronary artery spasm, evaluated by ST-segment elevation on lead II of body surface electrocardiogram, was induced by intravenous ergometrine injection (30mg/kg) in all controls (5/5, 100%) and all propranolol-treated mice (5/5, 100%), whereas it tended to be inhibited in carvedilol-treated mice (3/6, 50%) (p=0.08 vs control). In isolated VSMCs obtained from the aorta of AKAO-KO mice, the changes in [Ca2+]i from baseline in response to acetylcholine (ACh, 100μM) did not differ among the groups of pretreatment with carvedilol (10μM), propranolol (10μM), or vehicle, indicating that coronary artery spasm in AKAP-KO mice was induced by Ca2+-independent mechanism. In response to ACh, the phosphorylation of CaMK II was tended to be decreased (p=0.07), and the phosphorylation of myosin light chain was decreased (p<0.05) in VSMCs treated with carvedilol compared with control. Conclusions Carvedilol may inhibit coronary artery spasm probably via inhibiting CaMK II activity in AKAP-KO mice. Our results may provide a new paradigm into the pathogenesis of CSA as a model of enhanced Ca2+ sensitivity and important clinical implications for the mechanism of calcium channel antagonist-refractory coronary artery spasm.

The CNP analogue vosoritide reduces arrhythmia in diabetic cardiomyopathy via cGMP-dependent PDE2 stimulation on cellular and organ level

Abstract Background Detrimental arrhythmia occurs frequently in patients with diabetic cardiomyopathy (DiabCM), but safe and effective therapies are limited. Diabetes-associated neuropathological alterations increase the sympathetic tone, leading to hyperactivation of β-adrenoceptor-mediated cAMP signalling in cardiomyocytes (CM). Increased activity of deleterious cAMP-dependent kinases causes pro-arrhythmic dysregulation of intracellular Ca2+ homeostasis. Phosphodiesterases (PDE) can reduce cAMP levels. Exceptionally, the cAMP-hydrolysing activity of PDE2 can be cGMP-dependently increased by C-type natriuretic peptide (CNP). Thus, the CNP analogue vosoritide (approved for the treatment of achondroplasia) might mediate this cGMP/cAMP crosstalk, reducing arrhythmia. Purpose Here, we aim to study whether the cGMP-dependent activation of PDE2 by vosoritide (VO) is sufficient to reduce arrhythmia and arrhythmogenic triggers in DiabCM. Methods Diabetes was induced by 5 consecutive injections of streptozotocin (50 mg/kg) in control (WT) mice and mice with a CM-specific PDE2 knockout (KO). All experiments were performed 5 weeks later. Cardiac function was characterised by echocardiography. Primary ventricular CM were isolated to assess protein expression and phosphorylation levels by Western blot (WB) and cAMP levels by ELISA. Patch-clamp and Ca2+ imaging experiments were used to investigate pro-arrhythmic triggers. Arrhythmic events were quantified in ex vivo perfused hearts after ischaemia-reperfusion (I/R) injury. Results Compared to healthy controls, diabetic animals showed impaired systolic function with a ~9 % reduction in ejection fraction. The significant increase in E/E’ ratio by ~34 % indicated a diastolic dysfunction. Interestingly, the cardiac function was further worsened in diabetic PDE2 KO. WB analysis revealed upregulation of the CNP receptor NPR-B and PDE2 in diabetic CM and altered regulation of proteins involved in cAMP-dependent Ca2+ cycling such as protein kinase A, Ca2+/calmodulin-dependent kinase II and phospholamban. Interestingly, VO significantly reduced isoprenaline (Iso)-induced cAMP levels in diabetic CM. The effect of VO was prevented by specific PDE2 inhibition with BAY 60-7550 (BAY). Furthermore, VO was able to reduce the Iso-induced increase in L-type Ca2+ current, whereas BAY or genetic PDE2 deletion counteracted this effect. Accordingly, VO significantly decreased the Iso-triggered Ca2+ spark frequency by ~40 %. Concomitant PDE2 inhibition or PDE2 KO prevented this anti-arrhythmic effect. Finally, ECG analysis revealed that VO clearly reduced arrhythmia development after I/R in ex vivo perfused diabetic hearts via PDE2 activation. Conclusion Our data indicate that vosoritide-induced PDE2 stimulation may attenuate abnormal cardiac Ca2+ cycling and thereby reduce arrhythmia in diabetic hearts. Thus, vosoritide may be repurposed as a novel anti-arrhythmic drug in DiabCM.

Secretoneurin plasma levels are decreased after catheter ablation for atrial fibrillation, patients with AF produce lower SN levels than healthy individuals

Abstract Background Secretoneurin (SN) is a novel biomarker with potential use in cardiovascular medicine. The main effect of SN is mediated through its inhibition of calmodulin-dependent kinase II (CaMKII), which influences calcium handling[1], plasma SN levels differ in ischemic vs. dilated cardiomyopathy [2]. We aim to associate the levels of SN pre- and postprocedurally and according to the type of atrial fibrillation (AF). Methods We prospectively enrolled patients indicated for catheter ablation (CA) for AF, sequentially sampled patients for SN before, during and after CA for AF. We also sampled healthy individuals as controls. SN was analysed from venous, left atrial and coronary sinus (CS) blood by the use of the ELISA method. Values are expressed as means±SD, variables are compared by standard statistical methods. Results A total of 104 patients (59.4% male) with AF aged 64±10.4, with BMI 30.1±5 and 34 healthy individuals (38% male) were included in the analysis. SN plasma levels were significantly lower 32.3±15.1 after the procedure (SN2) as compared to levels before (SN1) 38.4±17.5 pmol/L (P=0.02, Fig.1). SN was not associated with any marker of cardiac injury or overload, echocardiographic parameters, or ablation characteristics (P=NS), even though Troponin I was significantly increased after the procedure (P<0.0001, Fig.2). Left atrial SN was numerically higher 40.9±19.1 than SN sampled from CS 32.9±13 (P=NS).The SN levels in our patients with AF were generally lower than in healthy individuals (50.7±15.3 pmol/L) before (P=0.0004) as well as after the procedure (P=0.0001). Conclusion Catheter ablation of AF leads to a decrease in SN plasma levels even though markers of cardiac injury are increased. Patients with AF irrespective of actual rhythm produce lower levels of SN than healthy individuals.

Functions and mechanisms of CDPKs in plant responses to abiotic stress

Calcium-dependent protein kinases (CDPKs/CPKs) are members of the Ca2+-sensitive Ser/Thr protein kinase family and play a crucial role in plant growth and development and responses to abiotic stress. CDPKs are capable of rapidly sensing changes in intracellular Ca2+ signals and recognizing and phosphorylating specific substrates, thereby transmitting and amplifying Ca2+ signal cascades downstream. They are involved in plant responses to stress conditions such as drought, saline-alkali stress, and injuries and regulate plant growth and development, gene expression, ion channel activity, and stomatal movement. The autophosphorylation of CDPKs can affect their activities and substrate specificity. CDPKs have the ability to bind to and phosphorylate multiple substrates. In addition to participating in respiratory burst oxidase homolog (RBOH), mitogen-activated protein kinase (MAPK), and plant hormone signaling pathways, CDPKs can also bind to 14-3-3 proteins, which enables the regulation of plant responses to stress and promotes plant growth and development. This paper summarized the research findings on the discovery, structure, classification, and roles of CDPKs in plant responses to stress and proposed the future research directions, aiming to provide the genetic resources and a theoretical basis for improving the stress tolerance of crops.

Subfamily C7 Raf-like kinases MRK1, RAF26, and RAF39 regulate immune homeostasis and stomatal opening in Arabidopsis thaliana.

The calcium-dependent protein kinase CPK28 regulates several stress pathways in multiple plant species. Here, we aimed to discover CPK28-associated proteins in Arabidopsis thaliana. We used affinity-based proteomics and identified several potential CPK28 binding partners, including the C7 Raf-like kinases MRK1, RAF26, and RAF39. We used biochemistry, genetics, and physiological assays to gain insight into their function. We define redundant roles for these kinases in stomatal opening, immune-triggered reactive oxygen species (ROS) production, and resistance to a bacterial pathogen. We report that CPK28 associates with and trans-phosphorylates RAF26 and RAF39, and that MRK1, RAF26, and RAF39 are active kinases that localize to endomembranes. Although Raf-like kinases share some features with mitogen-activated protein kinase kinase kinases (MKKKs), we found that MRK1, RAF26, and RAF39 are unable to trans-phosphorylate any of the 10 Arabidopsis mitogen-activated protein kinase kinases (MKKs). Overall, our study suggests that C7 Raf-like kinases associate with and are phosphorylated by CPK28, function redundantly in stomatal opening and immunity, and possess substrate specificities distinct from canonical MKKKs.

Heat stress causes chromatin accessibility and related gene expression changes in crown tissues of barley (Hordeum vulgare).

Plant responses to stress caused by high temperatures involve changes occurring at the molecular, metabolic, and physiological levels. Understanding the mechanisms by which plants recognize signals to activate this response is a prerequisite for identifying key genes and signaling pathways and for obtaining heat-tolerant plants. We demonstrated the first implementation of an assay for transposase-accessible chromatin to identify open chromatin regions (OCRs) in crown tissues of barley using three genotypes carrying different allelic forms of the sdw1 gene encoding gibberellin 20-oxidase subjected to elevated temperatures. In parallel, we performed gene expression analysis, which allowed us to relate changes in chromatin state to changes in transcriptional activity. The obtained data revealed that the hypersensitive chromatin regions within the genes were more repeatable than those outside the gene intervals. We observed that prolonged exposure to high temperatures increased chromatin accessibility. Genes with OCRs in their regulatory regions were involved in stress signaling and tolerance, including calcium-dependent protein kinase, mitogen-activated protein kinase (MAPK3), receptor-like cytoplasmic kinase (RLK), TIFY domain-containing transcriptional regulator, bZIP transcription factor, and regulatory protein NPR1. The effect of genotype on gene expression was not as pronounced as that of temperature. By combining results from the differential analysis of chromatin accessibility and expression profiles, we identified genes with high temperature-induced changes in chromatin accessibility associated with expression alterations. Importantly, our data revealed a relationship between the loss of chromatin accessibility in response to heat and the downregulation of genes related to gibberellin signaling.

Baicalein Ameliorates Insulin Resistance of HFD/STZ Mice Through Activating PI3K/AKT Signal Pathway of Liver and Skeletal Muscle in a GLP-1R-Dependent Manner.

Insulin resistance (IR) is the principal pathophysiological change occurring in diabetes mellitus (DM). Baicalein, a bioactive flavonoid primarily extracted from the medicinal plant Scutellaria baicalensis Georgi, has been shown in our previous research to be a potential natural glucagon-like peptide-1 receptor (GLP-1R) agonist. However, the exact therapeutic effect of baicalein on DM and its underlying mechanisms remain elusive. In this study, we investigated the therapeutic effects of baicalein on diabetes and sought to clarify its underlying molecular mechanisms. Our results demonstrated that baicalein improves hyperglycemic, hyperinsulinemic, and glucometabolic disorders in mice with induced diabetes via GLP-1R. This was confirmed by the finding that baicalein's effects on improving IR were largely diminished in mice with whole-body Glp1r ablation. Complementarily, network pharmacology analysis highlighted the pivotal involvement of the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) insulin signaling pathway in the therapeutic actions of baicalein on IR. Our mechanism research significantly confirmed that baicalein mitigates hepatic and muscular IR through the PI3K/AKT signal pathway, both in vitro and in vivo. Furthermore, we demonstrated that baicalein enhances glucose uptake in skeletal muscle cells under IR conditions through the Ca2+/calmodulin-dependent protein kinase II (CaMKII)-adenosine 5'-monophosphate-activated protein kinase (AMPK)-glucose transporter 4 (GLUT4) signaling pathway in a GLP-1R-dependent manner. In conclusion, our findings confirm the therapeutic effects of baicalein on IR and reveal that it improves IR in liver and muscle tissues through the PI3K/AKT insulin signaling pathway in a GLP-1R dependent manner. Moreover, we clarified that baicalein enhances the glucose uptake in skeletal muscle tissue through the Ca2+/CaMKII-AMPK-GLUT4 signal pathway.

Transcriptome sequencing analysis of overexpressed SikCDPK1 in tobacco reveals mechanisms of cold stress response

BackgroundCold is a significant limiting factor in productivity, particularly in northwestern and eastern China. Calcium-Dependent Protein Kinases (CDPKs), a primary calcium signaling sensor in plants, play an important role in their response to cold. Snow lotus (Sasussured involucrata Kar L) is a plant that thrives in harsh climates and grows in northwest China. However, there were no reports on the transcriptome of OE-SikCDPK1 transgenic tobacco in response to cold.ResultsWhen exposed to cold stress, OE-SikCDPK1 plants displayed a cold-tolerant phenotype compared to non-transgenic tobacco. Under cold conditions, relative water content reduced, relative conductivity increased, malondialdehyde levels rose, and cold-responsive gene expression increased. The OE-SikCDPK1 gene and non-transgenic tobacco were employed for research purposes. The transcriptome of leaves was sequenced using the HISAT2 sequencing platform, and the data were used to examine gene function annotation and differentially expressed genes (DEGs). 53,022 DEGs in tobacco leaves under cold treatment were obtained. The GO enrichment results revealed that it was enriched for biological-process, defense response and other processes under cold stress. The KEGG pathway enrichment analysis revealed that the metabolic pathways of significant enrichment of DEGs under cold stress mainly involved MAPK signaling pathway transduction. The transcription factor identification results showed that the transcription factors with the largest number of differential expressions under cold treatment were mainly from WRKY, AP2, MYB, bHLH, NAC and other transcription factor families.ConclusionThe cold tolerance mechanism of snow lotus SikCDPK1 was comprehensively analyzed at the transcriptional level for the first time using RNA-seq technology. This study demonstrates that SikCDPK1 can respond to cold by participating in the MAPK signaling pathway and regulating the expression levels of transcription factors, including WRKY, AP2, MYB, bHLH, and NAC. These results offer valuable insights for further exploration of the cold tolerance mechanism associated with SikCDPK1.

Extracellular RIPK3 Acts as a Danger-Associated Molecular Pattern to Exaggerate Cardiac Ischemia/Reperfusion Injury.

Cardiac ischemia/reperfusion (I/R) injury has emerged as an important therapeutic target for ischemic heart disease. Currently, there is no effective therapy for reducing cardiac I/R injury. Damage-associated molecular patterns are endogenous molecules released after cellular damage to exaggerate tissue inflammation and injury. RIPK3 (receptor-interacting protein kinase 3), a well-established intracellular mediator of cell necroptosis and inflammation, serves as a circulating biomarker of multiple diseases. However, whether extracellular RIPK3 also exerts biological functions in cardiac I/R injury remains totally unknown. Patients with acute myocardial infarction receiving percutaneous coronary intervention (PCI) were recruited independently in the discovery cohort (103 patients) and validation cohort (334 patients), and major adverse cardiovascular events were recorded. Plasma samples were collected before and after PCI (6 and 24 h) for RIPK3 concentration measurement. Cultured neonatal rat ventricular myocytes, macrophages and endothelial cells, and in vivo mouse models with myocardial injury induced by I/R (or hypoxia/reoxygenation) were used to investigate the role and mechanisms of extracellular RIPK3. Another cohort including patients with acute myocardial infarction receiving PCI and healthy volunteers was recruited to further explore the mechanisms of extracellular RIPK3. In the discovery cohort, elevated plasma RIPK3 levels after PCI are associated with poorer short- and long-term outcomes in patients with acute myocardial infarction, as confirmed in the validation cohort. In both cultured cells and in vivo mouse models, recombinant RIPK3 protein exaggerated myocardial I/R (or hypoxia/reoxygenation) injury, which was alleviated by the RIPK3 antibody. Mechanistically, RIPK3 acted as a damage-associated molecular pattern and bound with RAGE (receptor of advanced glycation end-products), subsequently activating CaMKII (Ca2+/calmodulin-dependent kinase II) to elicit the detrimental effects. The positive correlation between plasma RIPK3 concentrations and CaMKII phosphorylation in human peripheral blood mononuclear cells was confirmed. We identified the positive relationship between plasma RIPK3 concentrations and the risk of major adverse cardiovascular events in patients with acute myocardial infarction receiving PCI. As a damage-associated molecular pattern, extracellular RIPK3 plays a causal role in multiple pathological conditions during cardiac I/R injury through RAGE/CaMKII signaling. These findings expand our understanding of the physiological and pathological roles of RIPK3, and also provide a promising therapeutic target for myocardial I/R injury and the associated complications.

The Role of Human Adiponectin Receptor 1 in 2-Ethylhexyl Diphenyl Phosphate Induced Lipid Metabolic Disruption.

Epidemiological evidence links exposure to 2-ethylhexyl diphenyl phosphate (EHDPP) with lipid metabolic disruption, typically attributed to nuclear receptors, while the role of membrane receptors remains underexplored. This study explored the role of adiponectin receptor 1 (AdipoR1) in EHDPP-induced lipid metabolic disturbances. We examined EHDPP's binding affinity and transcriptional impact on AdipoR1. AdipoR1 knockdown (AdipoR1kd) human liver cells and coculture experiments with AdipoR1 activator (AdipoRon) were used to investigate the effect and the mechanism. EHDPP disrupted triglyceride and phospholipid synthesis and altered corresponding gene expression, mirroring effects in AdipoR1kd cells but diminishing in EHDPP-treated AdipoR1kd cells. RNA sequencing revealed that EHDPP primarily disrupted oxidative phosphorylation and insulin signaling dependent on AdipoR1. Mechanistically, EHDPP interacted with AdipoR1 and reduced AdipoR1 protein levels at 10-7 mol/L or higher, weakening the activation of the calmodulin dependent protein kinase β (CaMKKβ)/AMPK/acetyl CoA carboxylase pathway. Furthermore, EHDPP pretreatment blocked the increase in Ca2+ flux and the corresponding kinase CaMKKβ, as well as liver kinase B1 (LKB1) activation induced by AdipoRon, which is necessary for AMPK activation. Collectively, these findings demonstrate that EHDPP-induced lipid imbalance is partially dependent on AdipoR1, expanding the understanding of environmental metabolic disruptors beyond nuclear receptors.

Loss of mitochondrial Ca2+ response and CaMKII/ERK activation by LRRK2R1441G mutation correlate with impaired depolarization-induced mitophagy

BackgroundStress-induced activation of ERK/Drp1 serves as a checkpoint in the segregation of damaged mitochondria for autophagic clearance (mitophagy). Elevated cytosolic calcium (Ca2+) activates ERK, which is pivotal to mitophagy initiation. This process is altered in Parkinson’s disease (PD) with mutations in leucine-rich repeat kinase 2 (LRRK2), potentially contributing to mitochondrial dysfunction. Pathogenic LRRK2 mutation is linked to dysregulated cellular Ca2+ signaling but the mechanism involved remains unclear.MethodsMitochondrial damages lead to membrane depolarization. To investigate how LRRK2 mutation impairs cellular response to mitochondrial damages, mitochondrial depolarization was induced by artificial uncoupler (FCCP) in wild-type (WT) and LRRK2R1441G mutant knockin (KI) mouse embryonic fibroblasts (MEFs). The resultant cytosolic Ca2+ flux was assessed using live-cell Ca2+ imaging. The role of mitochondria in FCCP-induced cytosolic Ca2+ surge was confirmed by co-treatment with the mitochondrial sodium-calcium exchanger (NCLX) inhibitor. Cellular mitochondrial quality and function were evaluated by Seahorse™ real-time cell metabolic analysis, flow cytometry, and confocal imaging. Mitochondrial morphology was visualized using transmission electron microscopy (TEM). Activation (phosphorylation) of stress response pathways were assessed by immunoblotting.ResultsAcute mitochondrial depolarization induced by FCCP resulted in an immediate cytosolic Ca2+ surge in WT MEFs, mediated predominantly via mitochondrial NCLX. However, such cytosolic Ca2+ response was abolished in LRRK2 KI MEFs. This loss of response in KI was associated with impaired activation of Ca2+/calmodulin-dependent kinase II (CaMKII) and MEK, the two upstream kinases of ERK. Treatment of LRRK2 inhibitor did not rescue this phenotype indicating that it was not caused by mutant LRRK2 kinase hyperactivity. KI MEFs exhibited swollen mitochondria with distorted cristae, depolarized mitochondrial membrane potential, and reduced mitochondrial Ca2+ store and mitochondrial calcium uniporter (MCU) expression. These mutant cells also exhibited lower cellular ATP: ADP ratio albeit higher basal respiration than WT, indicating compensation for mitochondrial dysfunction. These defects may hinder cellular stress response and signals to Drp1-mediated mitophagy, as evident by impaired mitochondrial clearance in the mutant.ConclusionsPathogenic LRRK2R1441G mutation abolished mitochondrial depolarization-induced Ca2+ response and impaired the basal mitochondrial clearance. Inherent defects from LRRK2 mutation have weakened the cellular ability to scavenge damaged mitochondria, which may further aggravate mitochondrial dysfunction and neurodegeneration in PD.

Dendritic, delayed, stochastic CaMKII activation in behavioural time scale plasticity.

Behavioural time scale plasticity (BTSP) is non-Hebbian plasticity induced by integrating presynaptic and postsynaptic components separated by a behaviourally relevant time scale (seconds)1. BTSP in hippocampal CA1 neurons underlies place cell formation. However, the molecular mechanisms that enable synapse-specific plasticity on a behavioural time scale are unknown. Here we show that BTSP can be induced in a single dendritic spine using two-photon glutamate uncaging paired with postsynaptic current injection temporally separated by a behavioural time scale. Using an improved Ca2+/calmodulin-dependent kinase II (CaMKII) sensor, we did not detect CaMKII activation during this BTSP induction. Instead, we observed dendritic, delayed and stochastic CaMKII activation (DDSC) associated with Ca2+ influx and plateau potentials 10-100 s after BTSP induction. DDSC required both presynaptic and postsynaptic activity, which suggests that CaMKII can integrate these two signals. Also, optogenetically blocking CaMKII 15-30 s after the BTSP protocol inhibited synaptic potentiation, which indicated that DDSC is an essential mechanism of BTSP. IP3-dependent intracellular Ca2+ release facilitated both DDSC and BTSP. Thus, our study suggests that non-synapse-specific CaMKII activation provides an instructive signal with an extensive time window over tens of seconds during BTSP.

Regulation mechanism of exogenous nitric oxide on phenanthrene uptake by ryegrass roots

Polycyclic aromatic hydrocarbons (PAHs) constitute a category of persistent organic contaminants that possess the potential to induce carcinogenic, teratogenic, and mutagenic consequences. Our previous findings have revealed that plant roots actively take up PAHs through co-transport with protons, and auxin can promote PAHs uptake by wheat roots. It remains unclear whether nitric oxide (NO), a signaling molecule involved in numerous physiological processes in plants and downstream of auxin, can affect PAHs uptake by plant roots. In our study, 50 μmol/L sodium nitroprusside (SNP) significantly enhanced phenanthrene uptake after 4 h of exposure. After the addition of SNP (50 μmol/L), the H+ flux on root surface increased, and H+-ATPase activity was activated, indicating that exogenous NO promotes phenanthrene uptake by plant roots via activating H+-ATPase. By studying the effects of 50 μmol/L cyclic guanosine monophosphate (cGMP), 5 mmol/L Ca2+, and 50 μmol/L adenosine monophosphate (AMP) on phenanthrene uptake by ryegrass roots and measuring root calcium-dependent protein kinases (CDPK) activity, we demonstrated that exogenous NO promotes phenanthrene uptake through the signaling pathway of NO, cGMP, Ca2+, CDPK, 14-3-3 protein and H+-ATPase. The results contribute significant insights into elucidating the underlying mechanisms of NO modulating PAHs absorption by plant roots, thereby offering crucial strategies for advancing food safety measures and enhancing the phytoremediation potential of soils and waters contaminated with PAHs.

TaCDPK1-5A positively regulates drought response through modulating osmotic stress responsive-associated processes in wheat (Triticum aestivum).

Wheat TaCDPK1-5A plays critical roles in mediating drought tolerance through regulating osmotic stress-associated physiological processes. Calcium (Ca2+) acts as an essential second messenger in plant signaling pathways and impacts plant abiotic stress responses. This study reported the function of TaCDPK1-5A, a calcium-dependent protein kinase (CDPK) gene in T. aestivum, in mediating drought tolerance. TaCDPK1-5A sensitively responded to drought and exogenous abscisic acid (ABA) signaling, displaying induced transcripts in plants under drought and ABA treatments. Yeast two-hybrid and co-immunoprecipitation assays revealed that TaCDPK1-5A interacts with the mitogen-activated protein kinase TaMAPK4-7D whereas the latter with ABF transcription factor TaABF1-3A, suggesting that TaCDPK1-5A constitutes a signaling module with above partners to transduce signals initiated by drought/ABA stressors. Overexpression of TaCDPK1-5A, TaMAPK4-7D and TaABF1-3A enhanced plant drought adaptation by modulating the osmotic stress-related physiological indices, including increased osmolyte contents, enlarged root morphology, and promoted stomata closure. Yeast one-hybrid assays indicated the binding ability of TaABF1-3A with promoters of TaP5CS1-1B, TaPIN3-5A, and TaSLAC1-3-2A, the genes encoding P5CS enzyme, PIN-FORMED protein, and slow anion channel, respectively. ChIP-PCR and transcriptional activation assays confirmed that TaABF1-3A regulates these genes at transcriptional level. Moreover, transgene analysis indicated that these stress-responsive genes positively regulated proline biosynthesis (TaP5CS1-1B), root morphology (TaPIN3-5A), and stomata closing (TaSLAC1-3-2A) upon drought signaling. Positive correlations were observed between yield and the transcripts of TaCDPK1-5A signaling partners in wheat cultivars under drought condition, with haplotype TaCDPK1-5A-Hap1 contributing to improved drought tolerance. Our study concluded that TaCDPK1-5A positively regulates drought adaptation and is a valuable target for molecular breeding the drought-tolerant cultivars in T. aestivum.

Unravelling the molecular mechanism underlying drought stress tolerance in Dinanath (Pennisetum pedicellatum Trin.) grass via integrated transcriptomic and metabolomic analyses

Dinanath grass (Pennisetum pedicellatum Trin.) is an extensively grown forage grass known for its significant drought resilience. In order to comprehensively grasp the adaptive mechanism of Dinanath grass in response to water deficient conditions, transcriptomic and metabolomics were applied in the leaves of Dinanath grass exposed to two distinct drought intensities (48-hour and 96-hour). Transcriptomic analysis of Dinanath grass leaves revealed that a total of 218 and 704 genes were differentially expressed under 48- and 96-hour drought conditions, respectively. The genes that were expressed differently (DEGs) and the metabolites that accumulated in response to 48-hour drought stress mainly showed enrichment in the biosynthesis of secondary metabolites, particularly phenolics and flavonoids. Conversely, under 96-hour drought conditions, the enriched pathways predominantly involved lipid metabolism, specifically sterol lipids. In particular, phenylpropanoid pathway and brassinosteroid signaling played a crucial role in drought response to 48- and 96-hour water deficit conditions, respectively. This variation in drought response indicates that the adaptation mechanism in Dinanath grass is highly dependent on the intensity of drought stress. In addition, different genes associated with phenylpropanoid and fatty acid biosynthesis, as well as signal transduction pathways namely phenylalanine ammonia-lyase, putrescine hydroxycinnamoyl transferase, abscisic acid 8’-hydroxylase 2, syntaxin-61, lipoxygenase 5, calcium-dependent protein kinase and phospholipase D alpha one, positively regulated with drought tolerance. Combined transcriptomic and metabolomic analyses highlights the outstanding involvement of regulatory pathways related to secondary cell wall thickening and lignin biosynthesis in imparting drought tolerance to Dinanath grass leaves. These findings collectively contribute to an enhanced understanding of candidate genes and key metabolites relevant to drought response in Dinanath grass. Furthermore, they establish a groundwork for the creation of a transcriptome database aimed at developing abiotic stress-tolerant grasses and major crop varieties through both transgenic and genome editing approaches.

Calcium-Dependent Protein Kinase 5 and 13 enhance salt tolerance in rice by directly activating OsMPK3/6 kinases.

Mitogen-activated protein kinases (MAPKs/MPKs) are pivotal regulators in many stress-signaling pathways in plants. The dual phosphorylation of the TXY motif by MAP Kinase Kinases (MKKs) is essential for activating MAPKs. Here, we reveal a mechanism for MAPK activation that bypasses the need for MKKs. We identified rice (Oryza sativa) calcium-dependent protein kinase 5 (OsCPK5) and OsCPK13as positive regulators in salt stress tolerance. These kinases are essential for the full activation of OsMPK3 and OsMPK6 in response to elevated sodium levels, with both OsMPK3 and OsMPK6 also acting as positive regulators in rice salt tolerance. Biochemical analysis demonstrated that OsCPK5/13 directly interact with and activate OsMPK3/6 by phosphorylating the TXY motif in vitro and in vivo. Additionally, we have discovered that OsCPK5/13 relocate from the cell membrane to the nucleus in response to salt stress. This process relies on their N-terminal myristoylation and a calcium-dependent phosphorylation event within the N-terminus. Our results elucidate a MAPK activation pathway in rice that is independent of traditional MKK-mediated phosphorylation, highlighting the crucial roles of OsCPK5 and OsCPK13 in directly phosphorylating and activating OsMPK3/6, which are important for rice tolerance to salt stress.

Prevention and control of cardiac arrhythmic by using therapeutic foods: A review.

Arrhythmia occurs as a common heart vascular disease. Functional food is a rich source of natural compounds with significant pharmacological, The aim of this paper is to explore its effect on arrhythmia. By reviewing the literature and summarising the findings, we described the role of functional foods in the alleviation of cardiac arrhythmias from different perspectives. Our study shows that functional foods have anti-arrhythmic effects through modulation of ion channels, oxidative stress, and Calmodulin-dependent protein kinase II. We summarize the mechanism of arrhythmia inhibition by the active ingredients of medicinal diets in this review article, intending to provide research ideas for dietary therapy to regulate arrhythmia.

Early life seizures and olfactory communication in rats.

Early life seizures (ELS) are commonly associated with autism spectrum disorder (ASD); however, the exact role of ELS in the pathology is unknown. Prior studies have demonstrated social deficits, a core feature of ASD, following ELS; consequently, alterations in sensory modalities may contribute to the overall social deficits. Considering the speculated contribution of sensory deficit to social communication, we examined the developmental consequences of early postnatal kainic acid (KA)-induced seizures on olfactory preference and neural markers in the olfactory bulb in both male and female Sprague Dawley rats. KA-induced seizures or saline was administered. Rats were then exposed to a series of biologically relevant scents including male scent, female scent, nest scent, and phenylethylamine during the juvenile period and again during adulthood. Alterations in sensory modalities were expected to be expressed via abnormal preference for certain scents and/or production of abnormal ultrasonic vocalizations in response to scents. The olfactory bulbs were also assessed for the biologically relevant markers glial fibrillary acidic protein (GFAP) and calcium/calmodulin-dependent protein kinase II (CAMKII). Our findings resulted in no significant differences in olfactory preference following ELS for juveniles or adults compared to controls. Similarly, there were no differences in GFAP expression or the ratio of phosphorylated CAMKII to CAMKII in either olfactory bulb. Interestingly, despite a lack of treatment differences, different scents were shown to elicit different responses in juvenile rats, yet these differences subsided in adulthood. Overall, the results of this study suggest that olfaction does not contribute to socialization deficit following ELS within the KA model.

Calmodulin kinase II inhibition suppresses atrioventricular conduction by regulating intracellular Ca2+ homeostasis

Ca2+/calmodulin-dependent protein kinase II(CaMKII) inhibition decelerates atrioventricular node (AVN) conduction, providing a potential treatment for tachycardia. However, the effectiveness of CaMKII inhibition on tachycardia and its underlying mechanism remains unclear. To assess the effectiveness of CaMKII inhibition in reducing ventricular rates during atrial fibrillation and elucidate the underlying mechanism in affecting AVN electrophysiology. Cardiac CaMKII inhibition (AC3-I) mice were used. Transesophageal atrial pacing was performed to evaluate AVN conduction function and induce atrial fibrillation. Patch clamp techniques were employed to record action potentials and ionic currents in AVN cells. Intracellular Ca2+ transients and sarcomere length measurements were obtained using the IonOptix system. Masson trichrome stain was used to evaluate fibrosis in the AVN region. Western blotting and immunofluorescence techniques were employed to detect connexin expression and localization. CaMKII inhibition decreased the ventricular rate during atrial fibrillation and isoproterenol-induced tachycardia. Esophageal electrocardiogram results from AC3-I mice showed longer AVN conduction than wild-type (WT) mice. AN and N-type AVN cells from AC3-I mice exhibited slower action potential frequencies and diastolic depolarization rates (DRR) than those of WT mice. The study revealed that CaMKII inhibition reduced AVN cell sarcoplasmic reticulum (SR) Ca2+ content, Ca2+ release rate from the SR during diastole, Ca2+ transient amplitude, and SR Ca2+ uptake rate. Additionally, CaMKII inhibition prolonged the sarcomere diastole duration and enhanced the sensitivity of sarcomeres to Ca2+. CaMKII inhibition effectively decreases the ventricular rate during atrial fibrillation and tachycardia by slowing down AVN conduction through suppressing Ca2+ overload in AVN cells.

Dopamine and D1 receptor in hippocampal dentate gyrus involved in chronic stress-induced alteration of spatial learning and memory in rats

There is increasing evidence that chronic stress (CS), which occurs when the body is exposed to prolonged stressors, significantly impairs learning and memory. Dopamine (DA) plays a critical role in learning and memory in the hippocampus through the activation of D1-like receptors (D1R). However, the specific roles of DA and D1R in the hippocampal dentate gyrus (DG), particularly in relation to CS-induced changes in spatial learning and memory, are not well understood. In this study, we established a CS rat model through the random application of various stressors. We assessed spatial learning and memory using the Morris water maze (MWM) and measured DA concentration and the amplitude of field excitatory postsynaptic potentials (fEPSP) in the DG during the MWM test in freely moving rats. We also examined the involvement of D1R in spatial learning and memory by microinjecting its antagonist (SCH23390) into the DG, and then analyzed the expressions of phosphorylated (p-) Ca2+/calmodulin-dependent protein kinase II (CaMKII), protein kinase A (PKA), and cAMP-response element binding protein (CREB) in the DG using western blot. During the MWM test, compared with the control group, the escape latency was increased, and the percentage of distance in target quadrant and the number of platform crossings were decreased, in addition, the increase of fEPSP amplitude in the DG was significantly attenuated in CS group. In the control group, the DA concentration in the DG was significantly increased during the MWM test, and this response was enhanced in the CS group. Microinjection of SCH23390 into the DG significantly improved the spatial learning and memory impairments in CS rats, and reversed the inhibitory effect of CS on increase of fEPSP amplitude in the DG during the MWM test. Furthermore, SCH23390 partially reversed the inhibitory effects of CS on the expressions of p-CaMKII, p-PKA, and p-CREB in the DG. Our findings suggest that overactivation of the DA-D1R system in the hippocampal DG impairs spatial learning and memory and related synaptic plasticity in CS rats via downregulation of PKA-CREB signaling pathway.