Signaling Pathways In Response Research Articles

Potential Early Markers for Breast Cancer: A Proteomic Approach Comparing Saliva and Serum Samples in a Pilot Study.

Breast cancer is the second leading cause of death for women in the United States, and early detection could offer patients the opportunity to receive early intervention. The current methods of diagnosis rely on mammograms and have relatively high rates of false positivity, causing anxiety in patients. We sought to identify protein markers in saliva and serum for early detection of breast cancer. A rigorous analysis was performed for individual saliva and serum samples from women without breast disease, and women diagnosed with benign or malignant breast disease, using isobaric tags for relative and absolute quantitation (iTRAQ) technique, and employing a random effects model. A total of 591 and 371 proteins were identified in saliva and serum samples from the same individuals, respectively. The differentially expressed proteins were mainly involved in exocytosis, secretion, immune response, neutrophil-mediated immunity and cytokine-mediated signaling pathway. Using a network biology approach, significantly expressed proteins in both biological fluids were evaluated for protein-protein interaction networks and further analyzed for these being potential biomarkers in breast cancer diagnosis and prognosis. Our systems approach illustrates a feasible platform for investigating the responsive proteomic profile in benign and malignant breast disease using saliva and serum from the same women.

Protein-Ligand Interactions in Scarcity: The Stringent Response from Bacteria to Metazoa, and the Unanswered Questions.

The stringent response, originally identified in Escherichia coli as a signal that leads to reprogramming of gene expression under starvation or nutrient deprivation, is now recognized as ubiquitous in all bacteria, and also as part of a broader survival strategy in diverse, other stress conditions. Much of our insight into this phenomenon derives from the role of hyperphosphorylated guanosine derivatives (pppGpp, ppGpp, pGpp; guanosine penta-, tetra- and tri-phosphate, respectively) that are synthesized on starvation cues and act as messengers or alarmones. These molecules, collectively referred to here as (p)ppGpp, orchestrate a complex network of biochemical steps that eventually lead to the repression of stable RNA synthesis, growth, and cell division, while promoting amino acid biosynthesis, survival, persistence, and virulence. In this analytical review, we summarize the mechanism of the major signaling pathways in the stringent response, consisting of the synthesis of the (p)ppGpp, their interaction with RNA polymerase, and diverse factors of macromolecular biosynthesis, leading to differential inhibition and activation of specific promoters. We also briefly touch upon the recently reported stringent-like response in a few eukaryotes, which is a very disparate mechanism involving MESH1 (Metazoan SpoT Homolog 1), a cytosolic NADPH phosphatase. Lastly, using ppGpp as an example, we speculate on possible pathways of simultaneous evolution of alarmones and their multiple targets.

The transcription factor POPEYE negatively regulates the expression of bHLH Ib genes to maintain iron homeostasis.

Iron (Fe) is an essential trace element for plants. When suffering from Fe deficiency, plants modulate the expression of Fe deficiency-responsive genes to promote Fe uptake. POPEYE (PYE) is a key bHLH (basic helix-loop-helix) transcription factor involved in Fe homeostasis. However, the molecular mechanism of PYE regulating the Fe deficiency response remains elusive in Arabidopsis. We found that the overexpression of PYE attenuates the expression of Fe deficiency-responsive genes. PYE directly represses the transcription of bHLH Ib genes (bHLH38, bHLH39, bHLH100, and bHLH101) by associating with their promoters. Although PYE contains an ethylene response factor-associated amphiphilic repression (EAR) motif, it does not interact with the transcriptional co-repressors TOPLESS/TOPLESS-RELATED (TPL/TPRs). Sub-cellular localization analysis indicated that PYE localizes in both the cytoplasm and nucleus. PYE contains a nuclear export signal (NES) which is required for the cytoplasmic localization of PYE. Mutation of the NES amplifies the repression function of PYE, resulting in down-regulation of Fe deficiency-responsive genes. Co-expression assays indicated that three bHLH IVc members (bHLH104, bHLH105/ILR3, and bHLH115) facilitate the nuclear accumulation of PYE. Conversely, PYE indirectly represses the transcription activation ability of bHLH IVc. Additionally, PYE directly negatively regulates its own transcription. This study provides new insights into the Fe deficiency response signalling pathway and enhances the understanding of PYE functions in Arabidopsis.

A Proteomic Screen to Unravel the Molecular Pathways Associated with Warfarin-Induced or TNAP-Inhibited Arterial Calcification in Rats

Arterial media calcification refers to the pathological deposition of calcium phosphate crystals in the arterial wall. This pathology is a common and life-threatening complication in chronic kidney disease, diabetes and osteoporosis patients. Recently, we reported that the use of a TNAP inhibitor, SBI-425, attenuated arterial media calcification in a warfarin rat model. Employing a high-dimensionality unbiased proteomic approach, we also investigated the molecular signaling events associated with blocking arterial calcification through SBI-425 dosing. The remedial actions of SBI-425 were strongly associated with (i) a significant downregulation of inflammatory (acute phase response signaling) and steroid/glucose nuclear receptor signaling (LXR/RXR signaling) pathways and (ii) an upregulation of mitochondrial metabolic pathways (TCA cycle II and Fatty Acid β-oxidation I). Interestingly, we previously demonstrated that uremic toxin-induced arterial calcification contributes to the activation of the acute phase response signaling pathway. Therefore, both studies suggest a strong link between acute phase response signaling and arterial calcification across different conditions. The identification of therapeutic targets in these molecular signaling pathways may pave the way to novel therapies against the development of arterial media calcification.

Scaffold Hopping and Screening for Potent Small Molecule Agonists for GRP94: Implications to Alleviate ER Stress-Associated Pathogenesis.

Disparity in the activity of Endoplasmic reticulum (ER) leads to degenerative diseases, mainly associated with protein misfolding and aggregation leading to cellular dysfunction and damage, ultimately contributing to ER stress. ER stress activates the complex network of Unfolded Protein Response (UPR) signaling pathways mediated by transmembrane proteins IRE1, ATF6, and PERK. In addition to UPR, many ER chaperones have evolved to optimize the output of properly foldedsecretory and membrane proteins. Glucose-regulated protein 94 (GRP94), an ER chaperone of heat shock protein HSP90 family, directs protein folding through interaction with other components of the ER protein folding machinery and assists in ER-associated degradation (ERAD). Activation of GRP94 would increase the efficacy of protein folding machinery and regulate the UPR pathway toward homeostasis. The present study aims to screen for novel agonists for GRP94 based on Core hopping, pharmacophore hypothesis, 3D-QSAR, and virtual screening with small-molecule compound libraries in order to improve the efficiency of native protein folding by enhancing GRP94 chaperone activity, therefore to reduce protein misfolding and aggregation. In this study, we have employed the strategy of small molecule-dependent ER programming to enhance the chaperone activity of GRP94 through scaffold hopping-based screening approach to identify specific GRP94 agonists. New scaffolds generated by altering the cores of NECA, the known GRP94 agonist, were validated by employing pharmacophore hypothesis testing, 3D-QSAR modeling, and molecular dynamics simulations. This facilitated the identification of small molecules to improve the efficiency of native protein folding by enhancing GRP94 activity. High-throughput virtual screening of the selected pharmacophore hypothesis against Selleckchem and ZINC databases retrieved a total of 2,27,081 compounds. Further analysis on docking and ADMET properties revealed Epimedin A, Narcissoside, Eriocitrin 1,2,3,4,6-O-Pentagalloylglucose, Secoisolariciresinol diglucoside, ZINC92952357, ZINC67650204, and ZINC72457930 as potential lead molecules. The stability and interaction of these small molecules were far better than the known agonist, NECA indicating their efficacy in selectively alleviating ER stress-associated pathogenesis. These results substantiate the fact that small molecule-dependent ER reprogramming would activate the ER chaperones and therefore reduce the protein misfolding as well as aggregation associated with ER stress in order to restore cellular homeostasis.

CBL-Interacting Protein Kinases 18 (CIPK18) Gene Positively Regulates Drought Resistance in Potato.

Sensor-responder complexes comprising calcineurin B-like (CBL) proteins and CBL-interacting protein kinases (CIPKs) are plant-specific Ca2+ receptors, and the CBL-CIPK module is widely involved in plant growth and development and a large number of abiotic stress response signaling pathways. In this study, the potato cv. "Atlantic" was subjected to a water deficiency treatment and the expression of StCIPK18 gene was detected by qRT-PCR. The subcellular localization of StCIPK18 protein was observed by a confocal laser scanning microscope. The StCIPK18 interacting protein was identified and verified by yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC). StCIPK18 overexpression and StCIPK18 knockout plants were constructed. The phenotypic changes under drought stress were indicated by water loss rate, relative water content, MDA and proline contents, and CAT, SOD and POD activities. The results showed that StCIPK18 expression was upregulated under drought stress. StCIPK18 is localized in the cell membrane and cytoplasm. Y2H shows the interaction between StCIPK18 and StCBL1, StCBL4, StCBL6 and StCBL8. BiFC further verifies the reliability of the interaction between StCIPK18 and StCBL4. Under drought stress, StCIPK18 overexpression decreased the water loss rate and MDA, and increased RWC, proline contents and CAT, SOD and POD activities; however, StCIPK18 knockout showed opposite results, compared with the wild type, in response to drought stress. The results can provide information for the molecular mechanism of the StCIPK18 regulating potato response to drought stress.

EIF5A2 specifically regulates the transcription of aging-related genes in human neuroblastoma cells

BackgroundPost-transcriptional regulation plays a critical role in controlling biological processes such as aging. Previous studies have shown that eukaryotic initiation factor 5A (EIF5A) might play a crucial role in aging. It is unknown whether EIF5A2, a second isoform of EIF5A, could impact aging through post-transcriptional regulation.MethodsIn the present study, EIF5A2 overexpression (EIF5A2-OE) was induced in SH-SY5Y cells. RNA-seq, bioinformatics analysis and RT-qPCR validation experiments were then performed to explore the molecular mechanism of EIF5A2-mediated transcriptional regulation. Cell viability, proportion of senescent cells and the cell cycle were respectively determined by Cell Counting Kit-8, SA-β‑galactosidase and flow cytometry to evaluate the cell senescence.ResultsA total of 190 downregulated and 126 upregulated genes related to EIF5A2-OE were identified. Genes closely related to cellular aging processes such as unfolded protein response (UPR), cell adhesion and calcium signaling pathway were under global transcriptional regulation. Moreover, EIF5A2-OE promoted the viability of SH-SY5Y cells and reduced cell senescence in vitro. Among 30 genes with the most significant expression differences in EIF5A2-OE cells, we identified eight genes, including ASNS, ATF3, ATF4, CEBPB, DDIT3, HERPUD1, HSPA5 and XBP1, enriched in the UPR. Through EIF5A2-tanscription factors (TFs)-targets regulation network in EIF5A2-OE cells, we found three TFs, BHLHE40, RHOXF1 and TBX20, that targeted at these eight UPR-related genes. Verification test via the published database of human glial cell tissue showed only BHLHE40 and RHOXF1 were significantly associated with EIF5A2.ConclusionsOur findings suggest that EIF5A2 may alleviate cell senescence in vitro and mediate UPR-related genes via specific TFs. Thus, EIF5A2 could function as a regulator of aging via the regulation of transcription, which greatly expands the current understanding of the mechanisms of EIF5A2-mediated gene regulation.

Procyanidin B2 3,3″-di-O-gallate suppresses IFN-γ production in murine CD4+ T cells through the regulation of glutamine influx via direct interaction with ASCT2.

Excessive activation of CD4+ T cells increases cytokine production substantially and induces immune-mediated diseases. Procyanidins are polyphenols with anti-inflammatory properties. Procyanidin B2 (PCB2) gallate [specifically, PCB2 3,3''-di-O-gallate (PCB2DG)] inhibits cytokine production through the suppression of glycolysis via mammalian target of rapamycin (mTOR) in T cells. Several amino acids play critical roles in T cell activation, especially glutamine, which is important in mTOR signaling and interferon-γ (IFN-γ) production in CD4+ T cells. However, the mechanisms underlying the effects of PCB2DG, including its interaction partners, have yet to be clarified. In the present study, the mechanisms underlying the inhibitory effect of PCB2DG on IFN-γ through glutamine metabolism regulation were investigated. We found that PCB2DG treatment reduced intracellular glutamine levels in CD4+ T cells, whereas the addition of glutamine abrogated the inhibitory effects of PCB2DG on IFN-γ production. The PCB2DG-induced reduction in intracellular glutamine accumulation led to the upregulated expression of activating transcription factor 4, which was induced by the cytoprotective signaling pathway in the amino acid response. In addition, the mRNA and protein expression levels of alanine serine cysteine transporter 2 (ASCT2), a major glutamine transporter in CD4+ T cells, were not altered by PCB2DG treatment. Further analysis using a target identification strategy revealed that PCB2DG binds to ASCT2, suggesting that PCB2DG interacts directly with this major glutamine transporter to inhibit glutamine influx. Overall, this study indicates that ASCT2 is a novel target protein of a dietary polyphenol and provides new insights into the mechanism underlying the immunomodulatory effects of polyphenols.

In Vitro and Predictive Computational Toxicology Methods for the Neurotoxic Pesticide Amitraz and Its Metabolites

The Varroa destructor parasite is responsible for varroasis in honeybees worldwide, the most destructive disease among parasitic diseases. Thus, different insecticides/acaricides have been widely used within beehives to control these parasitic diseases. Namely, amitraz is the most used acaricide due to its high efficacy shown against Varroa destructor. However, pesticides used for beehive treatments could be incorporated into the honey and accumulate in other hive products. Hence, honeybee health and the impairment of the quality of honey caused by pesticides have gained more attention. Amitraz and its main metabolites, N-(2,4-dimethylphenyl) formamide (2,4-DMF) and 2,4-dimethylaniline (2,4-DMA), are known to be potent neurotoxicants. In this research, the cytotoxicity of amitraz and its metabolites has been assessed by MTT and PC assays in HepG2 cells. In addition, possible target receptors by in silico strategies have been surveyed. Results showed that amitraz was more cytotoxic than its metabolites. According to the in silico ADMEt assays, amitraz and its metabolites were predicted to be compounds that are able to pass the blood-brain barrier (BBB) and induce toxicity in the central and peripheral nervous systems. The main target class predicted for amitraz was the family of A G protein-coupled receptors that comprises responses to hormones and neurotransmitters. This affects, among other things, reproduction, development, locomotion, and feeding. Furthermore, amitraz and its metabolites were predicted as active compounds interacting with diverse receptors of the Tox21-nuclear receptor signaling and stress response pathways.

Polymers of the candidalysin peptide become membrane pores.

C. albicans infection requires the action of the virulence factor candidalysin (CL), which damages the plasma membrane of target human cells (Moyes Nature 2016). However, the molecular mechanism that CL uses to permeabilize membranes was unknown. We have employed complementary biophysical, modeling, microscopy, and cell biology methods to reveal that CL forms membrane pores using a unique molecular mechanism (Russell eLife 2022). Unexpectedly, we observed that CL readily assembles into linear polymers in solution. We propose that the basic structural unit in polymer formation is a CL oligomer, which is sequentially added into a string configuration that can close into a loop. Our data indicate that CL loops spontaneously insert into the membrane to become membrane pores. We identified a CL mutation (G4W) that inhibited the formation of polymers in solution and prevented formation of pores in synthetic lipid membrane systems. Studies in epithelial cells showed that G4W CL failed to activate the danger response signaling pathway, a hallmark of the pathogenic effect of CL. These results indicate that CL polymerization in solution is a necessary step for the damage of cellular membranes. Analysis of thousands of CL pores by atomic force microscopy revealed co-existence of simple depressions and complex pores decorated with protrusions. Imaging and modeling indicate that the two types of pores are formed by CL oligomers assembled into alternate orientations. We propose that this structural rearrangement represents a maturation mechanism that stabilizes pore formation . The data show that CL uses a previously unknown mechanism to damage membranes, whereby pre-assembly of CL loops in solution directly leads to formation of membrane pores. Our investigation not only unravels a new paradigm for the formation of membrane pores, but additionally identifies CL polymerization as a novel therapeutic target to treat candidiasis.

The reprogrammed mouse heart phosphoproteome in response to the chronic stress of markedly high cardiac-specific adenylyl cyclase type 8 overexpression.

An appropriate intensity and duration of increased AC-cAMP-PKA-Ca2+ signaling is the quintessential component of acute adaptation to cardiac stress. Our another work demonstrated the existence of an exquisite heart performance and protection profile in response to chronically increased cardiac AC-cAMP-PKA-Ca2+ signaling in young adult mice with cardiac-specific overexpression of adenylyl cyclase type 8 (AC8) (TGAC8). Here, we conducted an unbiased phosphoproteomics analysis to determine the role of altered protein phosphorylation in the chronic adaptive heart performance and protection profile of left ventricle (LV) of TGAC8 at 3-4 months of age. We identified 12,963 phospho-peptides and 3646 phosphoproteins, including 4990 upregulated phosphopeptides and 781 differentially regulated (741 Up and 40 Down) phosphoproteins in TGAC8 vs. WT. Based on differentially regulated phosphoproteins by genotype, we discovered the enrichment of numerous stress response canonical pathways within reprogrammed TGAC8 LV, and predicted upstream regulators and downstream functions. In addition to PKA, numerous kinases (83) and phosphatases (30) were hyper-phosphorylated in TGAC8 vs. WT. By combining the phosphoproteome with its proteome and with the previously published TGAC8 transcriptome, we determined which cardiac performance and adaptive protection profiles were coordinately regulated at post-translational modification (PTM) (phosphorylation), translational and transcriptional levels. To assess the impact of phosphorylation on mRNA transcription, we determined the phosphorylation states of transcriptional factors (TFs) that differed by genotype. Hyper-phosphorylated TFs in TGAC8 were associated with increased mRNA transcription, and immune responses and metabolism pathways. To determine the coordination of proteome and phosphoproteome in reprogramming the TGAC8 LV, we normalized its phosphoproteome to its proteome. These analyses indicated that many stress response signaling pathways were consistently enriched at transcriptional, translational and PTM levels, and that PI3K/AKT signaling was also activated in TGAC8 LV.

Quantification of biophysical properties of candidalysin, a fungal peptide toxin secreted by C. albicans during invasion.

Candida albicans fungus can cause invasive candidiasis. To become pathogenic and translocate across the intestinal epithelium C.abicans fungus needs to change its morphology from yeast to invasive filamentous. Virulence factor candidalysin secreted during fungal invasion, is found to be crucial in direct cell membrane damage, the stimulation of a danger response signalling pathway, and the activation of epithelial immunity. Due to its amphiphilic alpha-helical structure peptide has a high binding affinity to plasma membranes. With increasing accumulation on the membrane surfaces, the alpha-helix peptide starts penetration of the lipid bilayer until the plasma membrane is damaged. However, whether there is specific affinity to different membrane compositions is still unknown. Here we determined the concentrations of candidalysin required for membrane permeabilization of large unilamellar vesicles with various lipid compositions. Then, we used spectral imaging to assess the lipid packing changes in giant unilamellar vesicles of different lipid composition resembling human epithelial, bacterial and fungal membranes under the treatment of candidalysin. Solvatochromic polarity-sensitive dyes were used to label model membranes. Spectral imaging data allowed to calculate generalized polarization map of the labelled membrane and further investigate lipid packing during peptide binding.

Transcriptome and computational approaches highlighting the molecular regulation of Zacco platypus induced by mesocosm exposure to common disinfectant chlorine

Chlorine (Cl2) is a disinfectant often used in swimming pools and water treatment facilities. However, it is released into aquatic ecosystems, where it may harm non-targeted organisms. Here, we performed a mesocosm experiment exposing Zacco platypus (Z. platypus) to biocide Cl2 for 30 days (30 d) at two days' time points 15 days (15 d) and 30 d samples were collected. Here, Z. platypus was exposed to a sublethal concentration (0.1 mg/L) of Cl2, and comparative transcriptomics analyses were performed to determine their response mechanisms at the molecular level. According to RNA sequencing of the whole-body transcriptome, 860 and 1189 differentially expressed genes (DEGs) were identified from the 15 d and 30 d responses to Cl2, respectively. After enrichment analysis of GO (Gene Ontology) functions and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways, identified DEGs were demonstrated to be associated with a variety of functions, including “ion binding and transmembrane transporters”. Cl2 also induced oxidative stress in Z. platypus by increasing the levels of reactive oxygen species (ROS) while decreasing the catalase (CAT) content and the levels of solute carrier family 22 member 11 (slc22a11), Caspase-8 (casp-8), inducible nitric oxide synthase (NOS2), cytosolic phospholipase A2 gamma (PLA2G4). However, Z. platypus still allows recovery during stress suspension by increasing the expression levels of solute carrier family proteins. The GO and KEGG annotation results revealed that the expression of DEGs were related to the detoxification process, immune response, and antioxidant mechanism. Additionally, protein-protein interaction networks (PPI) and cytoHubba analyses identified sixteen hub genes and their interaction. These findings elucidate the regulation of various DEGs and signaling pathways in response to Cl2 exposure, which will improve our knowledge and laid foundation for further investigation of the toxicity of Cl2 to Z. platypus.

CCDC115 inhibits autophagy-mediated degradation of YAP to promote cell proliferation.

Autophagy and Hippo signalling pathways both play important roles in cell homeostasis and are often involved in tumourigenesis. However, the crosstalk between these two signal pathways in response to stress conditions, such as nutrient deficiency, is incompletely understood. Here, we show that vesicular localised coiled-coil domain containing 115 (CCDC115) inhibits autophagy as well as Hippo signalling pathway under starvation. Moreover, we show that CCDC115 interacts with the HOPS complex. This interaction competes with STX17, thus inhibiting the fusion of autophagosomes with lysosomes. Hence, CCDC115 inhibits the autophagic degradation of yes-associated protein (YAP), thereby promoting cell proliferation in nutrient-restricted situation.

Design, synthesis, and biological evaluation of indolin-2-one derivatives as novel cyclin-dependent protein kinase 8 (CDK8) inhibitors

Cyclin-dependent protein kinase 8 (CDK8) plays important roles in regulating fibrotic growth factors and inflammatory signaling pathways. Long-term chronic inflammation of the lungs can lead to idiopathic pulmonary fibrosis (IPF). Abnormal alveolar epithelial regeneration leads to the release of various fibrotic growth factors and the activation of inflammatory cells. CDK8 regulates profibrotic cytokines broadly implicated in the pathogenesis of fibrosis. Therefore, inhibition of CDK8 is considered a promising strategy for treating IPF. Here, CDK8 inhibitors were designed and optimized using a fragment-based drug design strategy. Testing results revealed that 71% of the synthesized compounds inhibited CDK8 activity better than the original compound E966–0530. Of these compounds, compound 4k exhibited the strongest CDK8 enzyme-inhibiting activity (IC50 =129 nM). Notably, it displayed a 13-fold increase in potency when compared to E966–0530. Experiments on toxicity and inhibition of epithelial-mesenchymal transition (EMT) protein expressions showed that compound 4k can inhibit EMT protein expressions, but with no significant cytotoxicity for alveolar epithelial cells. Compound 4k showed a potent inhibitory effect in cell migration assays. Furthermore, compound 4k significantly inhibited the phosphorylation of p-Smad3 and RNA Pol II, which are critical mediators in the fibrotic response signaling pathway. Compound 4k remarkably reduced TGF-β1-induced oxidative stress. The above results reveal optimized CDK8 inhibitors with potential use for IPF therapeutic treatment.

A Coculture of Enterobacter and Comamonas Species Reduces Cadmium Accumulation in Rice.

The accumulation of cadmium (Cd) in plants is strongly impacted by soil microbes, but its mechanism remains poorly understood. Here, we report the mechanism of reduced Cd accumulation in rice by coculture of Enterobacter and Comamonas species. In pot experiments, inoculation with the coculture decreased Cd content in rice grain and increased the amount of nonbioavailable Cd in Cd-spiked soils. Fluorescence in situ hybridization and scanning electron microscopy detection showed that the coculture colonized in the rhizosphere and rice root vascular tissue and intercellular space. Soil metagenomics data showed that the coculture increased the abundance of sulfate reduction and biofilm formation genes and related bacterial species. Moreover, the coculture increased the content of organic matter, available nitrogen, and potassium and increased the activities of arylsulfatase, β-galactosidase, phenoloxidase, arylamidase, urease, dehydrogenase, and peroxidase in soils. In subsequent rice transcriptomics assays, we found that the inoculation with coculture activated a hypersensitive response, defense-related induction, and mitogen-activated protein kinase signaling pathway in rice. Heterologous protein expression in yeast confirmed the function of four Cd-binding proteins (HIP28-1, HIP28-4, BCP2, and CID8), a Cd efflux protein (BCP1), and three Cd uptake proteins (COPT4, NRAM5, and HKT6) in rice. Succinic acid and phenylalanine were subsequently proved to inhibit rice divalent Cd [Cd(II)] uptake and activate Cd(II) efflux in rice roots. Thus, we propose a model that the coculture protects rice against Cd stress via Cd immobilization in soils and reducing Cd uptake in rice. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Posttreatment with dexmedetomidine aggravates LPS-induced myocardial dysfunction partly via activating cardiac endothelial α2A-AR in mice

BackgroundDexmedetomidine (DEX) administered before or at 30 min after sepsis induction was reported to alleviate septic cardiomyopathy in experimental models. However, sepsis is a life-threatening organ dysfunction due to infection-induced dysregulated host response, whether DEX treatment in the presence of organ dysfunction affects septic cardiomyopathy is unknown. This study investigated the effect of DEX posttreatment on septic cardiomyopathy. MethodsMale wild-type and α2A-adrenergic receptor (AR) knockout mice were exposed to lipopolysaccharide (LPS) or cecal ligation puncture (CLP), and cultured cardiac endothelial cells were used. Mouse survival, myocardial function, inflammatory response and related signaling pathways were determined. ResultsDEX treatment at 6, 9 h after LPS challenge significantly reduced survival rate of LPS-challenged mice, especially at 9 h. DEX administered at 9 h after LPS injection or CLP significantly reduced survival in LPS or CLP-induced sepsis in wild-type mice, but not in α2A-AR knockout mice. LPS treatment for 20 h decreased the left ventricle + dp/dt, increased myocardial interleukin (IL)-1β and IL-6 concentrations as well as cardiac endothelial tumor necrosis factor (TNF)-α, vascular cell adhesion molecule-1 (VCAM-1) and ICAM-1 expression, which were enhanced by DEX treated at 9 h after LPS injection in wild-type mice, but not in α2A-AR knockout mice. Furthermore, DEX posttreatment increased p38 phosphorylation, c-Fos nuclear translocation and VCAM-1 expression in LPS-treated cardiac endothelial cells, which were eliminated by α2A-AR knockout or PKC inhibitor. ConclusionsDEX posttreatment aggravates LPS-induced cardiac inflammation and myocardial dysfunction, at least in part, via activating cardiac endothelial α2A-AR-mediated PKC signal pathway.

Genistein Protects against Acetaldehyde-Induced Oxidative Stress and Hepatocyte Injury in Chronic Alcohol-Fed Mice

Alcohol-related liver disease (ALD) is one of the most prevalent forms of liver disease in the world. Acetaldehyde, an intermediate product of alcohol catabolism, is a cause of liver injury caused by alcohol. This study was designed to evaluate the protective role and mechanism(s) of genistein against acetaldehyde-induced liver injury in the pathological process of ALD. We found that genistein administration significantly ameliorated alcohol-induced hepatic steatosis, injury, and inflammation in mice. Genistein supplementation markedly reversed hepatic oxidative stress, endoplasmic reticulum stress, mitochondrial dysfunction, and hepatocellular apoptosis in both alcohol-fed mice liver and acetaldehyde-treated hepatocytes. The mechanistic experiments revealed that the restoration of genistein administration rescued heme oxygenase-1 (HO-1) reduction at both transcriptional and protein levels in either alcohol-fed mice liver or acetaldehyde-treated hepatocytes, and the beneficial aspects derived from genistein were abolished in antioxidase heme oxygenase-1 (HO-1)-deficient hepatocytes. Moreover, we confirmed that genistein administration-restored hepatic nuclear factor erythroid 2-related factor 2 (NRF2), a key transcriptional regulator of HO-1, was involved in the protective role of genistein in ALD. This study demonstrated that genistein ameliorated acetaldehyde-induced oxidative stress and liver injury by restoring the hepatic NRF2-HO-1 signaling pathway in response to chronic alcohol consumption. Therefore, genistein may serve as a potential therapeutic choice for the treatment of ALD.

Transcriptome analysis reveals the differential inflammatory effects between propofol and sevoflurane during lung cancer resection: a randomized pilot study

BackgroundPropofol and sevoflurane are two commonly used perioperative anesthetics. Some studies have found that these anesthetic drugs affect tumorigenesis. Previous studies have mostly focused on in vitro experiments, and the specimens collected were mainly peripheral body fluids, lacking direct evidence of the impact of anesthetic drugs on human tissues. This study aimed to elucidate the effects of propofol and sevoflurane on lung cancer using next-generation sequencing through an in vivo experiment.MethodsPatients were randomly assigned to a group receiving either propofol or sevoflurane during surgery. Then, the patients’ tumor and paired normal samples were collected and sequenced by next-generation sequencing. Differentially expressed genes (DEG) were analyzed by two statistical models, followed by cluster analysis, PCA, Gene Ontology, and KEGG pathway analysis. Candidate genes were confirmed by qRT–PCR.ResultsThe demographic data of the two study groups were not statistically significant. Through single-factor model analysis, 810 DEG in the propofol group and 508 DEG in the sevoflurane group were obtained. To better reflect the differential effects between propofol and sevoflurane while reducing the false-positive DEG, we used multifactor model analysis, which resulted in 124 DEG. In PCA and cluster analysis, four groups (propofol cancer group, propofol normal group, sevoflurane cancer group, sevoflurane normal group) were separated adequately, indicating the accuracy of the analysis. We chose seven significant pathways (cellular response to interleukin-1, chemokine-mediated signaling pathway, chemokine signaling pathway, cytokine–cytokine receptor interaction, inflammatory response, immune response, and TNF signaling pathway) for downstream analysis. Based on the pathway analysis, three candidate genes (CXCR1, CXCL8, and TNFAIP3) were chosen, and their qRT–PCR results were consistent with the sequencing results.ConclusionsThrough RNA-seq analysis, the effects of propofol and sevoflurane during lung cancer resection were different, mainly in inflammatory-related pathways, which might be possibly by targeting CXCL8.Trial registrationTrial registry number was ChiCTR1900026213.

SLAM-seq reveals early transcriptomic response mechanisms upon glutamine deprivation in Chinese hamster ovary cells.

Mammalian cells frequently encounter subtle perturbations during recombinant protein production. Identifying the genetic factors that govern the cellular stress response can facilitate targeted genetic engineering to obtain production cell lines that demonstrate a higher stress tolerance. To simulate nutrient stress, Chinese hamster ovary (CHO) cells were transferred into a glutamine(Q)-free medium and transcriptional dynamics using thiol(SH)-linked alkylation for the metabolic sequencing of RNA (SLAM-seq) along with standard RNA-seq of stressed and unstressed cells were investigated. The SLAM-seq method allows differentiation between actively transcribed, nascent mRNA, and total (previously present) mRNA in the sample, adding an additional, time-resolved layer to classic RNA-sequencing. The cells tackle amino acid (AA) limitation by inducing the integrated stress response (ISR) signaling pathway, reflected in Atf4 overexpression in the early hours post Q deprivation, leading to subsequent activation of its targets, Asns, Atf3, Ddit3, Eif4ebp1, Gpt2, Herpud1, Slc7a1, Slc7a11, Slc38a2, Trib3, and Vegfa. The GCN2-eIF2α-ATF4 pathway is confirmed by a significant halt in transcription of translation-related genes at 24 h post Q deprivation. The downregulation of lipid synthesis indicates the inhibition of the mTOR pathway, further confirmed by overexpression of Sesn2. Furthermore, SLAM-seq detects short-lived transcription factors, such as Egr1, that would have been missed in standard experimental designs with RNA-seq. Our results describe the successful establishment of SLAM-seq in CHO cells and therefore facilitate its future use in other scenarios where dynamic transcriptome profiling in CHO cells is essential.