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Journal of Animal Science and... Feb 2023Necroptosis and pyroptosis are newly identified forms of programmed cell death, which play a vital role in development of many gastrointestinal disorders. Although plant...
BACKGROUND
Necroptosis and pyroptosis are newly identified forms of programmed cell death, which play a vital role in development of many gastrointestinal disorders. Although plant polyphenols have been reported to protect intestinal health, it is still unclear whether there is a beneficial role of plant polyphenols in modulating necroptosis and pyroptosis in intestinal porcine epithelial cell line (IPEC-1) infected with enterotoxigenic Escherichia coli (ETEC) K88. This research was conducted to explore whether plant polyphenols including protocatechuic acid (PCA) and quercetin (Que), attenuated inflammation and injury of IPEC-1 caused by ETEC K88 through regulating necroptosis and pyroptosis signaling pathways.
METHODS
IPEC-1 cells were treated with PCA (40 μmol/L) or Que (10 μmol/L) in the presence or absence of ETEC K88.
RESULTS
PCA and Que decreased ETEC K88 adhesion and endotoxin level (P < 0.05) in cell supernatant. PCA and Que increased cell number (P < 0.001) and decreased lactate dehydrogenases (LDH) activity (P < 0.05) in cell supernatant after ETEC infection. PCA and Que improved transepithelial electrical resistance (TEER) (P < 0.001) and reduced fluorescein isothiocyanate-labeled dextran (FD4) flux (P < 0.001), and enhanced membrane protein abundance of occludin, claudin-1 and ZO-1 (P < 0.05), and rescued distribution of these tight junction proteins (P < 0.05) after ETEC infection. PCA and Que also declined cell necrosis ratio (P < 0.05). PCA and Que reduced mRNA abundance and concentration of tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and IL-8 (P < 0.001), and down-regulated gene expression of toll-like receptors 4 (TLR4) and its downstream signals (P < 0.001) after ETEC infection. PCA and Que down-regulated protein abundance of total receptor interacting protein kinase 1 (t-RIP1), phosphorylated-RIP1 (p-RIP1), p-RIP1/t-RIP1, t-RIP3, p-RIP3, mixed lineage kinase domain-like protein (MLKL), p-MLKL, dynamin- related protein 1 (DRP1), phosphoglycerate mutase 5 (PGAM5) and high mobility group box 1 (HMGB1) (P < 0.05) after ETEC infection. Moreover, PCA and Que reduced protein abundance of nod-like receptor protein 3 (NLRP3), nod-like receptors family CARD domain-containing protein 4 (NLRC4), apoptosis-associated speck-like protein containing a CARD (ASC), gasdermin D (GSDMD) and caspase-1 (P < 0.05) after ETEC infection.
CONCLUSIONS
In general, our data suggest that PCA and Que are capable of attenuating ETEC-caused intestinal inflammation and damage via inhibiting necroptosis and pyroptosis signaling pathways.
PubMed: 36721159
DOI: 10.1186/s40104-022-00816-x -
BMC Microbiology Jan 2023Typhoid fever is transmitted by ingestion of polluted water, contaminated food, and stool of typhoid-infected individuals, mostly in developing countries with poor...
Genomic landscape of the emerging XDR Salmonella Typhi for mining druggable targets clpP, hisH, folP and gpmI and screening of novel TCM inhibitors, molecular docking and simulation analyses.
Typhoid fever is transmitted by ingestion of polluted water, contaminated food, and stool of typhoid-infected individuals, mostly in developing countries with poor hygienic environments. To find novel therapeutic targets and inhibitors, We employed a subtractive genomics strategy towards Salmonella Typhi and the complete genomes of eight strains were primarily subjected to the EDGAR tool to predict the core genome (n = 3207). Human non-homology (n = 2450) was followed by essential genes identification (n = 37). The STRING database predicted maximum protein-protein interactions, followed by cellular localization. The virulent/immunogenic ability of predicted genes were checked to differentiate drug and vaccine targets. Furthermore, the 3D models of the identified putative proteins encoded by the respective genes were constructed and subjected to druggability analyses where only "highly druggable" proteins were selected for molecular docking and simulation analyses. The putative targets ATP-dependent CLP protease proteolytic subunit, Imidazole glycerol phosphate synthase hisH, 7,8-dihydropteroate synthase folP and 2,3-bisphosphoglycerate-independent phosphoglycerate mutase gpmI were screened against a drug-like library (n = 12,000) and top hits were selected based on H-bonds, RMSD and energy scores. Finally, the ADMET properties for novel inhibitors ZINC19340748, ZINC09319798, ZINC00494142, ZINC32918650 were optimized followed by binding free energy (MM/PBSA) calculation for ligand-receptor complexes. The findings of this work are expected to aid in expediting the identification of novel protein targets and inhibitors in combating typhoid Salmonellosis, in addition to the already existing therapies.
Topics: Humans; Anti-Bacterial Agents; Endopeptidase Clp; Genomics; Molecular Docking Simulation; Salmonella typhi; Typhoid Fever
PubMed: 36681806
DOI: 10.1186/s12866-023-02756-6 -
Microorganisms Jan 2023The development of Bacillus spore cores involves the accumulation of 3-phosphoglycerate (3PGA) during sporulation, following core acidification to ~6.4, and before...
Role of Spore Core Water Content and pH in the Accumulation and Utilization of Spores' Large 3-Phosphoglyceric Acid Depot, and the Crucial Role of This Depot in Generating ATP Early during Spore Germination.
The development of Bacillus spore cores involves the accumulation of 3-phosphoglycerate (3PGA) during sporulation, following core acidification to ~6.4, and before decreases in core water content occur due to Ca-dipicolinc acid (CaDPA) uptake. This core acidification inhibits phosphoglycerate mutase (PGM) at pH 6.4, allowing 3PGA accumulation, although PGM is active at pH 7.4. Spores’ 3PGA is stable for months at 4 °C and weeks at 37 °C. However, in wild-type spore germination, increases in core pH to 7.5−8 and in core water content upon CaDPA release and cortex peptidoglycan hydrolysis allow for rapid 3PGA catabolism, generating ATP; indeed, the earliest ATP generated following germination is from 3PGA catabolism. The current work found no 3PGA in those Bacillus subtilis spores that do not accumulate CaDPA during sporulation and have a core pH of ~7.4. The ATP production in the germination of 3PGA-less spores in a poor medium was minimal, and the germinated spores were >99% dead. However, the 3PGA-replete spores that germinated in the poor medium accumulated >30 times more ATP, and >70% of the germinated spores were found to be alive. These findings indicate why 3PGA accumulation during sporulation (and utilization during germination) in all the Firmicute spores studied can be crucial for spore revival due to the generation of essential ATP. The latter finding further suggests that targeting PGM activity during germination could be a novel way to minimize the damaging effects of spores.
PubMed: 36677488
DOI: 10.3390/microorganisms11010195 -
The inhibition of PGAM5 suppresses seizures in a kainate-induced epilepsy model mitophagy reduction.Frontiers in Molecular Neuroscience 2022Epilepsy is a common neurological disease, and excessive mitophagy is considered as one of the major triggers of epilepsy. Mitophagy is a crucial pathway affecting...
BACKGROUND
Epilepsy is a common neurological disease, and excessive mitophagy is considered as one of the major triggers of epilepsy. Mitophagy is a crucial pathway affecting reactive oxygen species. Phosphoglycerate mutase 5 (PGAM5) is a protein phosphatase present in mitochondria that regulates many biological processes including mitophagy and cell death. However, the mechanism of PGAM5 in epilepsy remains unclear. The purpose of the present study was to examine whether PGAM5 affects epilepsy through PTEN-induced putative kinase 1 (PINK1)-mediated mitophagy.
METHODS
After the knockdown of PGAM5 expression by the adeno-associated virus, an epilepsy model was created by kainic acid. Next, the seizure activity was recorded by local field potentials before evaluating the level of mitochondrial autophagy marker proteins. Lastly, the ultrastructure of mitochondria, neuronal damage and oxidative stress levels were further observed.
RESULTS
A higher PGAM5 level was found in epilepsy, and its cellular localization was in neurons. The interactions between PGAM5 and PINK1 in epilepsy were further found. After the knockdown of PGAM5, the level of PINK1 and light chain 3B was decreased and the expression of the translocase of the inner mitochondrial membrane 23 and translocase of the outer mitochondrial membrane 20 were both increased. Knockdown of PGAM5 also resulted in reduced neuronal damage, decreased malondialdehyde levels, decreased reactive oxygen species production and increased superoxide dismutase activity. In addition, the duration of spontaneous seizure-like events (SLEs), the number of SLEs and the time spent in SLEs were all reduced in the epilepsy model after inhibition of PGAM5 expression.
CONCLUSION
Inhibition of PGAM5 expression reduces seizures inhibiting PINK1-mediated mitophagy.
PubMed: 36618822
DOI: 10.3389/fnmol.2022.1047801 -
Frontiers in Cell and Developmental... 2022Phosphoglycerate mutase 2 (PGAM2) is a critical glycolytic enzyme that is highly expressed in skeletal muscle. In humans, naturally occurring mutations in...
Phosphoglycerate mutase 2 (PGAM2) is a critical glycolytic enzyme that is highly expressed in skeletal muscle. In humans, naturally occurring mutations in Phosphoglycerate mutase 2 have been etiologically linked to glycogen storage disease X (GSDX). Phosphoglycerate mutase 2 activity is regulated by several posttranslational modifications such as ubiquitination and acetylation. Here, we report that Phosphoglycerate mutase 2 activity is regulated by sumoylation-a covalent conjugation involved in a wide spectrum of cellular events. We found that Phosphoglycerate mutase 2 contains two primary SUMO acceptor sites, lysine (K)49 and K176, and that the mutation of either K to arginine (R) abolished Phosphoglycerate mutase 2 sumoylation. Given that K176 is more highly evolutionarily conserved across paralogs and orthologs than K49 is, we used the CRISPR-mediated homologous recombination technique in myogenic C2C12 cells to generate homozygous K176R knock-in cells (PGAM2). Compared with wild-type (WT) C2C12 cells, PGAM2 C2C12 cells exhibited impaired myogenic differentiation, as indicated by decreased differentiation and fusion indexes. Furthermore, the results of glycolytic and mitochondrial stress assays with the XF96 Extracellular Flux analyzer revealed a reduced proton efflux rate (PER), glycolytic PER (glycoPER), extracellular acidification rate (ECAR), and oxygen consumption rate (OCR) in PGAM2 C2C12 cells, both at baseline and in response to stress. Impaired mitochondrial function was also observed in PGAM2 P19 cells, a carcinoma cell line. These findings indicate that the PGAM2-K176R mutation impaired glycolysis and mitochondrial function. Gene ontology term analysis of RNA sequencing data further revealed that several downregulated genes in PGAM2 C2C12 cells were associated with muscle differentiation/development/contraction programs. Finally, PGAM2 with either of two naturally occurring missense mutations linked to GSDX, E89A (conversion of glutamic acid 89 to alanine) or R90W (conversion of arginine 90 to tryptophan), exhibited reduced Phosphoglycerate mutase 2 sumoylation. Thus, sumoylation is an important mechanism that mediates Phosphoglycerate mutase 2 activity and is potentially implicated in Phosphoglycerate mutase 2 mutation-linked disease in humans.
PubMed: 36589741
DOI: 10.3389/fcell.2022.1052363 -
Frontiers in Plant Science 2022() is a gram-negative bacterium that causes bacterial fruit blotch (BFB) disease in cucurbit crops including watermelon. However, despite the great economic losses...
A putative 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase is involved in the virulence, carbohydrate metabolism, biofilm formation, twitching halo, and osmotic tolerance in .
() is a gram-negative bacterium that causes bacterial fruit blotch (BFB) disease in cucurbit crops including watermelon. However, despite the great economic losses caused by this disease worldwide, -resistant watermelon cultivars have not been developed. Therefore, characterizing the virulence factors/mechanisms of would enable the development of effective control strategies against BFB disease. The 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase (BdpM) is known to participate in the glycolysis and gluconeogenesis pathways. However, the roles of the protein have not been characterized in . To elucidate the functions of BdpmAc (Bdpm in ), comparative proteomic analysis and diverse phenotypic assays were conducted using a knockout mutant () and a wild-type strain. The virulence of the mutant to watermelon was remarkably reduced in both germinated seed inoculation and leaf infiltration assays. Moreover, the mutant could not grow with fructose or pyruvate as a sole carbon source. However, the growth of the mutant was restored to levels similar to those of the wild-type strain in the presence of both fructose and pyruvate. Comparative proteomic analyses revealed that diverse proteins involved in motility and wall/membrane/envelop biogenesis were differentially abundant. Furthermore, the mutant exhibited decreased biofilm formation and twitching halo size. Interestingly, the mutant exhibited a higher tolerance against osmotic stress. Overall, our findings suggest that BdpmAc affects the virulence, glycolysis/gluconeogenesis, biofilm formation, twitching halo size, and osmotic tolerance of , suggesting that this protein has pleiotropic properties. Collectively, our findings provide fundamental insights into the functions of a previously uncharacterized phosphoglycerate mutase in .
PubMed: 36438092
DOI: 10.3389/fpls.2022.1039420 -
Frontiers in Aging Neuroscience 2022We aimed to examine whether plasma-derived phosphoglycerate mutase 5 (PGAM5) can be a biomarker for Parkinson's disease (PD) diagnosis as well as its association with...
BACKGROUND
We aimed to examine whether plasma-derived phosphoglycerate mutase 5 (PGAM5) can be a biomarker for Parkinson's disease (PD) diagnosis as well as its association with the severity of motor/non-motor manifestations of PD.
METHODS
We enrolled 124 patients with PD (PD group) and 50 healthy controls (HC group). We measured plasma PGAM5 levels using a quantitative sandwich enzyme immunoassay. Patients with PD underwent baseline evaluations using the Unified Parkinson's Disease Rating Scale (UPDRS), while participants in both groups were evaluated using scales for non-motor manifestations. Receiver operating characteristic curves were used to evaluate the predictive utility of plasma PAMG5 alone and combined with other factors.
RESULTS
Plasma PAMG5 levels were significantly higher in the PD group; the area under the curve (AUC) of plasma PGAM5 levels alone was 0.76. The AUC values for elderly participants and patients without hypertension were 0.78 and that for was 0.79. Notably, plasma PGAM5 levels combined with plasma oligomeric α-synuclein (α-syn) and the score of the REM sleep behavior disorder questionnaire-Hong Kong (RBDQ-HK) showed AUC values of 0.80 and 0.82. Multivariable logistic analysis revealed that plasma PAMG5 levels were independently associated with PD (odds ratio,1.875 [95% confidence interval 1.206-2.916], = 0.005) but not the severity of motor/non-motor manifestations of PD.
CONCLUSION
Plasma PGAM5 is an independent biomarker for PD, especially among elderly patients (age > 60 years) and patients without hypertension. The predictive utility of PGAM5 was improved when combined with plasma oligomeric α-syn or the RBDQ-HK score.
PubMed: 36389083
DOI: 10.3389/fnagi.2022.1022274 -
International Journal of Molecular... Oct 2022Phosphoglycerate mutase (PGAM) is a glycolytic enzyme converting 3-phosphoglycerate to 2-phosphoglycerate, which in mammalian cells is expressed in two isoforms: brain...
Phosphoglycerate mutase (PGAM) is a glycolytic enzyme converting 3-phosphoglycerate to 2-phosphoglycerate, which in mammalian cells is expressed in two isoforms: brain (PGAM1) and muscle (PGAM2). Recently, it was shown that besides its enzymatic function, PGAM2 can be imported to the cell nucleus where it co-localizes with the nucleoli. It was suggested that it functions there to stabilize the nucleolar structure, maintain mRNA expression, and assist in the assembly of new pre-ribosomal subunits. However, the precise mechanism by which the protein translocates to the nucleus is unknown. In this study, we present the first crystal structure of PGAM2, identify the residues involved in the nuclear localization of the protein and propose that PGAM contains a "quaternary nuclear localization sequence (NLS)", i.e., one that consists of residues from different protein chains. Additionally, we identify potential interaction partners for PGAM2 in the nucleoli and demonstrate that 14-3-3ζ/δ is indeed an interaction partner of PGAM2 in the nucleus. We also present evidence that the insulin/IGF1-PI3K-Akt-mTOR signaling pathway is responsible for the nuclear localization of PGAM2.
Topics: Animals; Phosphoglycerate Mutase; Active Transport, Cell Nucleus; Phosphatidylinositol 3-Kinases; 14-3-3 Proteins; Muscles; Mammals
PubMed: 36361985
DOI: 10.3390/ijms232113198 -
Toxicology Letters Jan 2023The pathophysiology of renal lipid toxicity caused by excess adiposity is not well-understood. Necroptosis, a regulated form of cell death, is involved in injuring renal...
The pathophysiology of renal lipid toxicity caused by excess adiposity is not well-understood. Necroptosis, a regulated form of cell death, is involved in injuring renal tubular epithelial cells (RTECs). Phosphoglycerate mutase 5 (PGAM5) is a key downstream effector of necroptosis. This study investigated the underlying mechanism of PGAM5 in promoting lipid-induced necroptosis in RTECs. HK2 cells (an immortalized proximal tubule epithelial cell line) were exposed to oleic acid (OA) to mimic the lipid overload environment in vitro. We found that OA suppressed HK2 cell proliferation, triggered cytoskeleton rupture and cell death. In OA-treated cells, upregulated expression of necroptosis pathway proteins, phosphorylated receptor-interacting protein-1/3 (pRIPK1/3), phosphorylated mixed lineage kinase domain-like protein (pMLKL), PGAM5, phosphorylated dynamin-related protein 1 (pDRP1), and downregulated pDRP1 expression were observed. This was accompanied by mitochondrial dysfunction (mitochondrial ROS overproduction and decreased mitochondrial membrane potential) and increased cellular necrosis, as reflected by Annexin V/ Propidium Iodide (PI) labeling. OA also induced the accumulation of LC3II and P62, blocking autophagosome fusion with lysosomes. Knockdown of PGAM5 could prevent these OA-induced changes. We propose inhibition of PGAM5 protects lipid-induced RTECs from necroptosis by reducing DRP1-mediated mitochondrial fission and improving mitophagy flux.
Topics: Mitophagy; Mitochondrial Dynamics; Necroptosis; Phosphoglycerate Mutase; Epithelial Cells; Lipids; Mitochondrial Proteins
PubMed: 36273635
DOI: 10.1016/j.toxlet.2022.10.003 -
International Journal of Molecular... Sep 2022Stressors cause activation of the hypothalamic-pituitary-adrenal (HPA) axis and a systemic inflammatory response. As a newly proposed cell death manner in recent years,...
Stressors cause activation of the hypothalamic-pituitary-adrenal (HPA) axis and a systemic inflammatory response. As a newly proposed cell death manner in recent years, necroptosis occurs in a variety of tissue damage and inflammation. However, the role of necroptosis in HPA axis activation remains to be elucidated. The aim of this study was to investigate the occurrence of necroptosis and its role in HPA activation in a porcine stress model induced by lipopolysaccharide (LPS). Several typical stress behaviors like fever, anorexia, shivering and vomiting were observed in piglets after LPS injection. HPA axis was activated as shown by increased plasma cortisol concentration and mRNA expression of pituitary corticotropin-releasing hormone receptor 1 () and adrenal steroidogenic acute regulatory protein (). The mRNA expression of tumor necrosis factor α (), interleukin-1β () and in the hypothalamus, pituitary gland and adrenal gland was elevated by LPS, accompanied by the activation of necroptosis indicated by higher mRNA expression of necroptosis signals including receptor-interacting protein kinase (RIP) 1, RIP3, and phosphorylated mixed-lineage kinase domain-like protein (MLKL). Furthermore, necrostatin-1 (Nec-1), an inhibitor of necroptosis, inhibited necroptosis indicated by decreased mRNA levels of , , , and phosphoglycerate mutase family member 5 () in the hypothalamus, pituitary gland and adrenal gland. Nec-1 also decreased the mRNA expression of and and inhibited the activation of the HPA axis indicated by lower plasma cortisol concentration and mRNA expression of adrenal type 2 melanocortin receptor () and . These findings suggest that necroptosis is present and contributes to HPA axis activation induced by LPS. These findings provide a potential possibility for necroptosis as an intervention target for alleviating HPA axis activation and stress responses.
Topics: Animals; Corticotropin-Releasing Hormone; Hydrocortisone; Hypothalamo-Hypophyseal System; Interleukin-1beta; Interleukin-6; Lipopolysaccharides; Necroptosis; Phosphoglycerate Mutase; Pituitary-Adrenal System; Protein Kinases; RNA, Messenger; Swine; Tumor Necrosis Factor-alpha
PubMed: 36232518
DOI: 10.3390/ijms231911218