Phosphoglycerate Kinase Research Articles

Metabolism of hydrogen peroxide by Lactobacillus plantarum NJAU-01: A proteomics study.

This study aimed to investigate the time-course effect of Lactobacillus plantarum NJAU-01 in scavenging exogenous hydrogen peroxide (H2O2). The results showed that L. plantarum NJAU-01 at 107CFU/mL was able to eliminate a maximum of 4mM H2O2 within a prolonged lag phase and resume to proliferate during the following culture. Redox state in the start-lag phase (0h, without the addition of H2O2), indicated by glutathione and protein sulfhydryl, was impaired in the lag phase (3h and 12h) and then gradually recovered during subsequent growing stages (20h and 30h). By using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and proteomics analysis, a total of 163 proteins such as PhoP family transcriptional regulator, glutamine synthetase, peptide methionine sulfoxide reductase, thioredoxin reductase, ribosomal proteins, acetolactate synthase, ATP binding subunit ClpX, phosphoglycerate kinase, UvrABC system protein A and UvrABC system protein B were identified as differential proteins across the entire growth phase. Those proteins were mainly involved in H2O2 sensing, protein synthesis, repairing proteins and DNA lesions, amino sugar and nucleotide sugar metabolism. Our data suggest that biomolecules of L. plantarum NJAU-01 are oxidized to passively consume H2O2 and are restored by the enhanced protein and/or gene repair systems.

Study on the Key Genes and Molecular Mechanisms of IL-27 Promoting Keratinocytes Proliferation Based on Transcriptome Sequencing.

IL-27 involves psoriasis pathogenesis potentially by promoting excessive keratinocyte proliferation. However, the underlying mechanisms remain unclear. This study aims to explore the key genes and molecular mechanisms of IL-27-induced keratinocyte proliferation. Primary keratinocytes and immortalized human keratinocyte HaCaT cells were treated with different concentrations of IL-27 for 24 h and 48 h respectively. CCK-8 assay was used to detect cell viability and Western blot was used to detect the expression of CyclinE and CyclinB1. Primary keratinocytes and HaCaT cells were treated with IL-27, and their differentially expressed (DE) genes were obtained by transcriptome sequencing. Then Kyoto Encyclopedia of Genes and Genomes enrichment analysis was performed to predict related pathways, and the long non-coding RNA-microRNA-messenger RNA network and protein-protein interaction network were constructed to screen key genes. Biochemical experiments were performed to assess the content of glucose (Glu), lactic acid (LA), and ATP. Flow cytometry and Mito-Tracker Green staining were used to detect mitochondrial membrane potential and the number of mitochondria respectively. Western blot was performed to assess the expression of glucose transporter 1 (GLUT1), hexokinase 2 (HK2), lactate dehydrogenase A (LDHA), phosphoglycerate kinase 1 (PGK1), phosphorylated dynamin-related protein 1 (p-DRP1) (s637) and mitofusin 2 (MFN2). IL-27 concentration-dependently increased keratinocyte viability and the expression of CyclinE and CyclinB1. Bioinformatics analysis showed that the enriched pathways of DE genes were closely associated with cellular metabolism. miR-7-5p, EGFR, PRKCB, PLCB1 and CALM3 were key genes. IL-27 increased the content of LA, mitochondrial membrane potential, and the expression of GLUT1, HK2, LDHA, PGK1, p-DRP1 (s637), and MFN2, accompanied by decreased contents of Glu and ATP (P<0.001). IL-27 potentially promotes keratinocyte proliferation by enhancing glycolysis, mitochondrial function, and mitochondrial fusion. The findings of this study may be conducive to revealing the role of IL-27 in the pathogenesis of psoriasis.

Activation of Pgk1 Results in Reduced Protein Aggregation in Diverse Neurodegenerative Conditions.

The prevention of protein condensates has emerged as a new drug target to treat diverse neurodegenerative disorders. We previously reported that terazosin (TZ), a prescribed antagonist of the α1 adrenergic receptor, is an activator of phosphoglycerate kinase 1 (Pgk1) and Hsp90. In this study, we aimed to determine whether TZ prevents the formation of diverse pathological condensates in cell cultures and animal disease models. In primary neuron culture, TZ treatment reduced both the protein density and abundance of fused in sarcoma (FUS)-P525L-GFP, a disease-associated mutant form of FUS. Regarding the mechanism, we found that increased intracellular ATP levels were critical for the reduction in protein aggregate density. In addition, Hsp90 activation by TZ enhanced Hsp90 interaction with ULK1, a master regulator of autophagy. Through in vivo studies, we examined neuron-specific overexpression of tau in Drosophila, mouse models of APP/PS1 Alzheimer's disease (AD), and a rat model of multiple system atrophy (MSA) via the viral expression of α-synuclein in the striatum. TZ prevented and reversed the formation of pathological protein condensates. Together, our results suggest that activation of Pgk1 in cytosol may dissolve pathological protein aggregates via increased ATP levels and degrade these proteins via autophagy; the FUS-P525L degradation pathway in nucleus is unclear.

Modeling and simulation of single droplet drying in an acoustic levitator

We present a mathematical model for the full drying process of a single protein formulation droplet taking into account the convective impact arising from its levitation by a standing ultrasound wave. Using the finite element method allows us to compute the evaporation rate directly from the fully resolved heat and mass transfer within and around the levitated droplet. We apply our model to simulate the drying kinetics of pure water and aqueous phosphoglycerate kinase (PGK) droplets under various levitation and drying conditions. Empirical data from the literature are used to validate and discuss our numerical results. The acoustic streaming turns out to accelerate not only the first but also the second drying stage. Moreover, the dehydration of the protein molecules is found to be primarily responsible for their enzymatic inactivation throughout the drying process.

20-Hydroxyecdysone counteracts insulin to promote programmed cell death by modifying phosphoglycerate kinase 1

BackgroundThe regulation of glycolysis and autophagy during feeding and metamorphosis in holometabolous insects is a complex process that is not yet fully understood. Insulin regulates glycolysis during the larval feeding stage, allowing the insects to grow and live. However, during metamorphosis, 20-hydroxyecdysone (20E) takes over and regulates programmed cell death (PCD) in larval tissues, leading to degradation and ultimately enabling the insects to transform into adults. The precise mechanism through which these seemingly contradictory processes are coordinated remains unclear and requires further research. To understand the coordination of glycolysis and autophagy during development, we focused our investigation on the role of 20E and insulin in the regulation of phosphoglycerate kinase 1 (PGK1). We examined the glycolytic substrates and products, PGK1 glycolytic activity, and the posttranslational modification of PGK1 during the development of Helicoverpa armigera from feeding to metamorphosis.ResultsOur findings suggest that the coordination of glycolysis and autophagy during holometabolous insect development is regulated by a balance between 20E and insulin signaling pathways. Glycolysis and PGK1 expression levels were decreased during metamorphosis under the regulation of 20E. Insulin promoted glycolysis and cell proliferation via PGK1 phosphorylation, while 20E dephosphorylated PGK1 via phosphatase and tensin homolog (PTEN) to repress glycolysis. The phosphorylation of PGK1 at Y194 by insulin and its subsequent promotion of glycolysis and cell proliferation were important for tissue growth and differentiation during the feeding stage. However, during metamorphosis, the acetylation of PGK1 by 20E was key in initiating PCD. Knockdown of phosphorylated PGK1 by RNA interference (RNAi) at the feeding stage led to glycolysis suppression and small pupae. Insulin via histone deacetylase 3 (HDAC3) deacetylated PGK1, whereas 20E via acetyltransferase arrest-defective protein 1 (ARD1) induced PGK1 acetylation at K386 to stimulate PCD. Knockdown of acetylated-PGK1 by RNAi at the metamorphic stages led to PCD repression and delayed pupation.ConclusionsThe posttranslational modification of PGK1 determines its functions in cell proliferation and PCD. Insulin and 20E counteractively regulate PGK1 phosphorylation and acetylation to give it dual functions in cell proliferation and PCD.

NET Proteome in Established Type 1 Diabetes Is Enriched in Metabolic Proteins.

Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by a T-cell-mediated destruction of the pancreatic insulin-producing beta cells. A growing body of evidence suggests that abnormalities in neutrophils and neutrophil extracellular trap (NET) formation (NETosis) are associated with T1D pathophysiology. However, little information is available on whether these changes are primary neutrophil defects or related to the environmental signals encountered during active disease. In the present work, the NET proteome (NETome) of phorbol 12-myristate 13-acetate (PMA)- and ionomycin-stimulated neutrophils from people with established T1D compared to healthy controls (HC) was studied by proteomic analysis. Levels of NETosis, in addition to plasma levels of pro-inflammatory cytokines and NET markers, were comparable between T1D and HC subjects. However, the T1D NETome was distinct from that of HC in response to both stimuli. Quantitative analysis revealed that the T1D NETome was enriched in proteins belonging to metabolic pathways (i.e., phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase, and UTP-glucose-1-phosphate uridylyltransferase). Complementary metabolic profiling revealed that the rate of extracellular acidification, an approximate measure for glycolysis, and mitochondrial respiration were similar between T1D and HC neutrophils in response to both stimuli. The NETome of people with established T1D was enriched in metabolic proteins without an apparent alteration in the bio-energetic profile or dysregulated NETosis. This may reflect an adaptation mechanism employed by activated T1D neutrophils to avoid impaired glycolysis and consequently excessive or suboptimal NETosis, pivotal in innate immune defence and the resolution of inflammation.

Senecavirus A-induced glycolysis facilitates virus replication by promoting lactate production that attenuates the interaction between MAVS and RIG-I

Senecavirus A (SVA)-induced porcine idiopathic vesicular disease has caused huge economic losses worldwide. Glucose metabolism in the host cell is essential for SVA proliferation; however, the impact of the virus on glucose metabolism in host cells and the subsequent effects are still unknown. Here, glycolysis induced by SVA is shown to facilitate virus replication by promoting lactate production, which then attenuates the interaction between the mitochondrial antiviral-signaling protein (MAVS) and retinoic acid-inducible gene I (RIG-I). SVA induces glycolysis in PK-15 cells, as indicated by significantly increased expression of hexokinase 2 (HK2), 6-phosphofructokinase (PFKM), pyruvate kinase M (PKM), phosphoglycerate kinase 1 (PGK1), hypoxia-inducible factor-1 alpha (HIF-1α), and superoxide dismutase-2 (SOD2) in a dose-and replication-dependent manner, and enhanced lactate production, while reducing ATP generation. Overexpression of PKM, PGK1, HIF-1α, and PDK3 in PK-15 cells and high glucose concentrations promote SVA replication, while glycolytic inhibitors decrease it. Inhibition of RLR signaling allowed better replication of SVA by promoting lactate production to attenuate the interaction between MAVS and RIG-I, and regulatory effect of glycolysis on replication of SVA was mainly via RIG-I signaling. SVA infection in mice increased expression of PKM and PGK1 in tissues and serum yields of lactate. Mice treated with high glucose and administered sodium lactate showed elevated lactate levels and better SVA replication, as well as suppressed induction of RIG-I, interferon beta (IFNβ), IFNα, interferon-stimulated gene 15 (ISG15), and interleukin 6 (IL-6). The inhibitory effect on interferons was lower in mice administered sodium oxamate and low glucose compared to the high glucose, indicating that RLR signaling was inhibited by SVA infection through lactate in vitro and in vivo. These results provide a new perspective on the relationship between metabolism and innate immunity of the host in SVA infection, suggesting that glycolysis or lactate may be new targets against the virus.

Comparative transcriptomics discovers the genetic basis of contrasting waterlogging tolerance between two cultivated jute species

Farmers prefer dark jute (Corchorus olitorius var. O-9897) to white jute (Corchorus capsularis var. CVL-1) because dark jute produces more bast fiber and has greater fiber strength. Crossing incompatibility between these two jute species is the main genetic barrier to transfer waterlogging-tolerant capability from white jute to dark jute. So, to develop a waterlogging-tolerant dark jute variety, gene(s) of interest for certain morphological, anatomical, and physiological features must be transferred from white jute to dark jute without the use of traditional crossing techniques. Therefore, we investigated the comparative gene expressions of CVL-1 and O-9897 tap roots using RNA-Seq and qRT-PCR techniques as well as observed the morphological, histological and physiological behavior under waterlogging treatments. CVL-1 develops schizogenous aerenchyma for oxygen storage, as well as vascular tissues in its adventitious roots for moving water and nutrients from the water solution to the stem, which keeps its tap root less active and so saves more energy in its tap roots. O-9897, on the other hand, degrades its adventitious roots with lysigenous aerenchyma, which results in the formation of no vascular tissues. Thus, O-9897 solely uses its adventitious roots for oxygen storage, while its tap roots are used for water and nutrient transfer, resulting in constant energy depletion in O-9897 tap roots. Consequently, CVL-1 can maintain its normal growth and O-9897 diminishes its bast fiber yield with the lack of energy. The key genes involved in the biogenesis of energy and phytohormones have been more inducing in CVL-1 than O-9897. ATP-producing glycolytic enzyme, phosphoglycerate kinase (CCACVL1_10615) showed remarkable upregulation in CVL-1 while missing in O-9897, playing a vital role in CVL-1 waterlogging tolerance. Our findings reveal a novel mechanism of waterlogging tolerance in white jute, which can be applied to the development of waterlogging-tolerant dark jute varieties in the future.

Low nitrogen stress-induced transcriptome changes revealed the molecular response and tolerance characteristics in maintaining the C/N balance of sugar beet (Beta vulgaris L.).

Nitrogen (N) is an essential macronutrient for plants, acting as a common limiting factor for crop yield. The application of nitrogen fertilizer is related to the sustainable development of both crops and the environment. To further explore the molecular response of sugar beet under low nitrogen (LN) supply, transcriptome analysis was performed on the LN-tolerant germplasm '780016B/12 superior'. In total, 580 differentially expressed genes (DEGs) were identified in leaves, and 1,075 DEGs were identified in roots (log2 |FC| ≥ 1; q value < 0.05). Gene Ontology (GO), protein-protein interaction (PPI), and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses clarified the role and relationship of DEGs under LN stress. Most of the downregulated DEGs were closely related to "photosynthesis" and the metabolism of "photosynthesis-antenna proteins", "carbon", "nitrogen", and "glutathione", while the upregulated DEGs were involved in flavonoid and phenylalanine biosynthesis. For example, GLUDB (glutamate dehydrogenase B) was identified as a key downregulated gene, linking carbon, nitrogen, and glutamate metabolism. Thus, low nitrogen-tolerant sugar beet reduced energy expenditure mainly by reducing the synthesis of energy-consuming amino acids, which in turn improved tolerance to low nitrogen stress. The glutathione metabolism biosynthesis pathway was promoted to quench reactive oxygen species (ROS) and protect cells from oxidative damage. The expression levels of nitrogen assimilation and amino acid transport genes, such as NRT2.5 (high-affinity nitrate transporter), NR (nitrate reductase [NADH]), NIR (ferredoxin-nitrite reductase), GS (glutamine synthetase leaf isozyme), GLUDB, GST (glutathione transferase) and GGT3 (glutathione hydrolase 3) at low nitrogen levels play a decisive role in nitrogen utilization and may affect the conversion of the carbon skeleton. DFRA (dihydroflavonol 4-reductase) in roots was negatively correlated with NIR in leaves (coefficient = -0.98, p < 0.05), suggesting that there may be corresponding remote regulation between "flavonoid biosynthesis" and "nitrogen metabolism" in roots and leaves. FBP (fructose 1,6-bisphosphatase) and PGK (phosphoglycerate kinase) were significantly positively correlated (p < 0.001) with Ci (intercellular CO2 concentration). The reliability and reproducibility of the RNA-seq data were further confirmed by real-time fluorescence quantitative PCR (qRT-PCR) validation of 22 genes (R2 = 0.98). This study reveals possible pivotal genes and metabolic pathways for sugar beet adaptation to nitrogen-deficient environments.

Identification and evaluation of novel serum autoantibody biomarkers for early diagnosis of gastric cancer and precancerous lesion.

Early diagnosis is crucial for optimal prognosis of gastric cancer (GC). Hereby, we aimed to identify novel serum autoantibody-based biomarkers for precancerous lesion (PL) and early GC. We performed serological proteome analysis (SERPA) combined with nanoliter-liquid chromatography combined with quadrupole time of flight tandem mass spectrometry (Nano-LC-Q-TOF-MS/MS) to screen for GC-associated autoantibodies. The identified autoantibodies were analyzed for potential detection value for PL and GC by enzyme-linked immunosorbent assay (ELISA). Receiver operating characteristic (ROC) curves analysis was conducted to evaluate the accuracy of the biomarkers. We identified seven candidates, such as mRNA export factor (RAE1), Nucleophosmin 1 (NPM1), phosphoglycerate kinase 1 (PGK1), and ADP-ribosylation factor 4 (ARF4). Antibodies against all seven proteins were present at higher levels in sera from 242 patients (51 PL, 78 early GC, 113 advanced GC) compared with sera from 122 healthy individuals. RAE1-specific autoantibody discriminated best between patients at different GC stages, with area under the curve (AUC) values of 0.710, 0.745, and 0.804 for PL, early GC, and advanced GC, respectively. Two predictive models composed of gender, RAE1, PGK1, NPM1, and ARF4 autoantibodies (Model 2 for PL) and of age, gender, RAE1, PGK1, and NPM1 autoantibodies (Model 3 for early GC) had improved diagnostic efficiencies, with AUCs of 0.803 and 0.857, sensitivities of 66.7% and 75.6%, and specificities of 78.7% and 87.7%, respectively. The identified serum tumor-associated autoantibodies (TAAbs) may have good potential for early detection of GC and PL.

Abstract 4802: Identification of critical hypoxia induced factors in castrate resistant prostate cancer

Abstract Background: Prostate cancer (PCa) is the second most diagnosed cancer in men and the second cause of cancer-related death amongst men worldwide. PCa is a heterogeneous disease, and the outcome is worse for patients when the disease progresses from localized PCa to a castrate-resistant disease. Androgen receptor (AR) signaling is crucial for PCa development and has been a major therapeutic target for decades. Despite the success made with hormone therapy to block AR signaling, castration resistant disease can arise through mutations and amplifications of the androgen receptor (AR) and AR splice variants (AR-Vs). Amongst them, AR-V7 and AR-V12 can promote AR signaling independent of androgen. Furthermore, prostate tumors are highly hypoxic due to abnormal tumor vasculature. Hypoxia influences the efficacy of local and systemic treatment strategies, such as radiotherapy, hormone therapy, and chemotherapy, resulting in poorer clinical outcomes for PCa patients. Therefore, a complete understanding of AR and AR variant function, as well as how they are modulated by hypoxia and therapy is warranted for developing new strategies to treat PCa. Methods: To identify the transcriptional apparatus associated with the AR and its variants under normal and hypoxic conditions, we carried out Bio-ID proximity labeling mass spectrometry (BioID-MS). We generated stable androgen-independent human prostate cancer cell lines, PC-3 and DU145, expressing AR and the AR-V7 and AR-V12 variants fused to a MiniTurboID (MTID) enzyme. To delineate the AR proximal interaction network, we treated cells with androgen or vehicle for 3 hours and affinity-purified biotinylated proteins after the addition of biotin for the final 2 hours. Cells were grown in normoxia (20% O2) or treated with hypoxia (95% N2,5% CO2 and 0.5% O2) for 24 h. Nonspecific interactions were filtered using controls and statistical methods to identify high-confidence interactomes. Results: BioID proximity labeling in PC3 cells identified several AR-associated proteins, including many knowninteractors, validating the approach. Hypoxic conditions led to a striking alteration in proximal interactions and many metabolic proteins were highly enriched with AR upon exposure of cells to hypoxia for 24h. Among these, phosphoglycerate kinase 1 (PGK1) was a top hit. PGK1 has been reported to interact with AR to mediate the expression of genes that regulates cell proliferation and apoptosis. Additionally, PGK1 has an essential role in metabolic reprogramming induced by c-MYC and HIF-1α, leading to enhanced tumorigenesis. Conclusion: Bio-ID mass spectrometry was used to map the proximal interactome of AR and its variants, identifying novel interactions partners for further analysis. We found that hypoxia strongly alters interactors with the AR, and we identified PGK1 as a hypoxia induced AR interaction in PCA. Current work to functionally validate this interaction will be presented. Citation Format: Oloruntoba I. Osagie, Jordann Smakk, Deborah E. Citrin, Travis H. Stracker. Identification of critical hypoxia induced factors in castrate resistant prostate cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4802.

E2F regulation of the Phosphoglycerate kinase gene is functionally important in Drosophila development

The canonical role of the transcription factor E2F is to control the expression of cell cycle genes by binding to the E2F sites in their promoters. However, the list of putative E2F target genes is extensive and includes many metabolic genes, yet the significance of E2F in controlling the expression of these genes remains largely unknown. Here, we used the CRISPR/Cas9 technology to introduce point mutations in the E2F sites upstream of five endogenous metabolic genes in Drosophila melanogaster. We found that the impact of these mutations on both the recruitment of E2F and the expression of the target genes varied, with the glycolytic gene, Phosphoglycerate kinase (Pgk), being mostly affected. The loss of E2F regulation on the Pgk gene led to a decrease in glycolytic flux, tricarboxylic acidcycle intermediates levels, adenosine triphosphate (ATP) content, and an abnormal mitochondrial morphology. Remarkably, chromatin accessibility was significantly reduced at multiple genomic regions in PgkΔE2F mutants. These regions contained hundreds of genes, including metabolic genes that were downregulated in PgkΔE2F mutants. Moreover, PgkΔE2F animals had shortened life span and exhibited defects in high-energy consuming organs, such as ovaries and muscles. Collectively, our results illustrate how the pleiotropic effects on metabolism, gene expression, and development in the PgkΔE2F animals underscore the importance of E2F regulation on a single E2F target, Pgk.

Lysine Acetylome of Breast Cancer-Derived Small Extracellular Vesicles Reveals Specific Acetylation Patterns for Metabolic Enzymes

Cancer-derived small extracellular vesicles have been proposed as promising potential biomarkers for diagnosis and prognosis of breast cancer (BC). We performed a proteomic study of lysine acetylation of breast cancer-derived small extracellular vesicles (sEVs) to understand the potential role of the aberrant acetylated proteins in the biology of invasive ductal carcinoma and triple-negative BC. Three cell lines were used as models for this study: MCF10A (non-metastatic), MCF7 (estrogen and progesterone receptor-positive, metastatic) and MDA-MB-231 (triple-negative, highly metastatic). For a comprehensive protein acetylation analysis of the sEVs derived from each cell line, acetylated peptides were enriched using the anti-acetyl-lysine antibody, followed by LC-MS/MS analysis. In total, there were 118 lysine-acetylated peptides, of which 22, 58 and 82 have been identified in MCF10A, MCF7 and MDA-MB-231 cell lines, respectively. These acetylated peptides were mapped to 60 distinct proteins and mainly identified proteins involved in metabolic pathways. Among the acetylated proteins identified in cancer-derived sEVs from MCF7 and MDA-MB-231 cell lines are proteins associated with the glycolysis pathway, annexins and histones. Five acetylated enzymes from the glycolytic pathway, present only in cancer-derived sEVs, were validated. These include aldolase (ALDOA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK1), enolase (ENO) and pyruvate kinase M1/2 (PKM). For three of these enzymes (ALDOA, PGK1 and ENO) the specific enzymatic activity was significantly higher in MDA-MB-231 when compared with MCF10A-derived sEVs. This study reveals that sEVs contain acetylated glycolytic metabolic enzymes that could be interesting potential candidates for early BC diagnostics.

Predatory Strategies of Myxococcus xanthus: Prey Susceptibility to OMVs and Moonlighting Enzymes.

Predatory outer membrane vesicles (OMVs) secreted by myxobacteria fuse readily with the outer membranes of Gram-negative bacteria, introducing toxic cargo into their prey. Here we used a strain of the myxobacterium Myxococcus xanthus that produces fluorescent OMVs to assay the uptake of OMVs by a panel of Gram-negative bacteria. M. xanthus strains took up significantly less OMV material than the tested prey strains, suggesting that re-fusion of OMVs with producing organisms is somehow inhibited. The OMV killing activity against different prey correlated strongly with the predatory activity of myxobacterial cells, however, there was no correlation between OMV killing activity and their propensity to fuse with different prey. It has previously been proposed that M. xanthus GAPDH stimulates the predatory activity of OMVs by enhancing OMV fusion with prey cells. Therefore, we expressed and purified active fusion proteins of M. xanthus glyceraldehyde-3-phosphate dehydrogenase and phosphoglycerate kinase (GAPDH and PGK; moonlighting enzymes with additional activities beyond their roles in glycolysis/gluconeogenesis) to investigate any involvement in OMV-mediated predation. Neither GAPDH nor PGK caused lysis of prey cells or enhanced OMV-mediated lysis of prey cells. However, both enzymes were found to inhibit the growth of Escherichia coli, even in the absence of OMVs. Our results suggest that fusion efficiency is not a determinant of prey killing, but instead resistance to the cargo of OMVs and co-secreted enzymes dictates whether organisms can be preyed upon by myxobacteria.

Integrative proteogenomic characterization of early esophageal cancer

Esophageal squamous cell carcinoma (ESCC) is malignant while the carcinogenesis is still unclear. Here, we perform a comprehensive multi-omics analysis of 786 trace-tumor-samples from 154 ESCC patients, covering 9 histopathological stages and 3 phases. Proteogenomics elucidates cancer-driving waves in ESCC progression, and reveals the molecular characterization of alcohol drinking habit associated signatures. We discover chromosome 3q gain functions in the transmit from nontumor to intraepithelial neoplasia phases, and find TP53 mutation enhances DNA replication in intraepithelial neoplasia phase. The mutations of AKAP9 and MCAF1 upregulate glycolysis and Wnt signaling, respectively, in advanced-stage ESCC phase. Six major tracks related to different clinical features during ESCC progression are identified, which is validated by an independent cohort with another 256 samples. Hyperphosphorylated phosphoglycerate kinase 1 (PGK1, S203) is considered as a drug target in ESCC progression. This study provides insight into the understanding of ESCC molecular mechanism and the development of therapeutic targets.

LncRNA HOTTIP impacts the proliferation and differentiation of fibroblast-like synoviocytes in ankylosing spondylitis through the microRNA-30b-3p/PGK1 axis

ObjectiveLong non-coding RNAs (lncRNAs) and microRNAs (miRNAs) have been reported to exert regulatory effects on biological processes. This study intended to assess the role of the lncRNA HOXA transcript at the distal tip (HOTTIP)/miR-30b-3p/phosphoglycerate kinase 1 (PGK1) axis in ankylosing spondylitis (AS).MethodsLevels of HOTTIP, miR-30b-3p and PGK1 in AS synovial tissues and cultured AS fibroblast-like synoviocytes (ASFLSs) were assessed. The ASFLSs were identified and, respectively, treated with altered expression of HOTTIP and miR-30b-3p, and then, the proliferation and differentiation of the ASFLSs were assessed. The AS mouse models were established by injection of proteoglycan and Freund’s complete adjuvant and then were treated with altered expression of HOTTIP and miR-30b-3p, and the pathological changes and apoptosis of synoviocytes in mice’ synovial tissues were measured. The relationship of HOTTIP, miR-30b-3p and PGK1 was verified.ResultsHOTTIP and PGK1 were elevated, while miR-30b-3p was reduced in AS synovial tissues and ASFLSs. Elevated miR-30b-3p or inhibited HOTTIP restrained proliferation and differentiation of ASFLSs and also improved the pathological changes and promoted apoptosis of synoviocytes in mice’s synovial tissues. PGK1 was a target of miR-30b-3p, and miR-30b-3p could directly bind to HOTTIP. Silencing miR-30b-3p or overexpressing PGK1 reversed the improvement of AS by knocking down HOTTIP or up-regulating miR-30b-3p.ConclusionOur study suggests that reduced HOTTIP ameliorates AS progression by suppressing the proliferation and differentiation of ASFLSs through the interaction of miR-30b-3p and PGK1.

Circular RNA circHMCU promotes breast tumorigenesis through miR-4458/PGK1 regulatory cascade

BackgroundCircular RNAs (circRNAs) are abnormally expressed in breast cancer (BC). However, the biological function and mechanism of circHMCU still need to be further explored.MethodsThe expression levels of circHMCU, miR-4458 and phosphoglycerate kinase 1 (PGK1) were measured by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot. The glucose uptake, lactate production and ATP level were assayed by related commercial kits. Cell Counting Kit-8 (CCK8), 5’-ethynyl-2’-deoxyuridine (EdU) and flow cytometry assays were used to test cell proliferation and apoptosis, respectively. The migratory and invasive abilities were detected by transwell and wound-healing assays. The relationships among circHMCU, miR-4458 and PGK1 were verified by dual-luciferase reporter assay. The function of circHMCU in tumor growth was evaluated by animal studies.ResultsCircHMCU was upregulated in BC tissues and cell lines, whereas miR-4458 was downregulated. For biological experiments, circHMCU knockdown inhibited cell proliferation, migration, glycolysis, while promoted cell apoptosis. CircHMCU bound miR-4458, and miR-4458 targeted PGK1. MiR-4458 inhibition reversed circHMCM knockdown-mediated effects on BC cell malignant behaviors. MiR-4458 overexpression suppressed cell glycolysis, proliferation, and metastasis and promoted apoptosis in BC cells through PGK1 upregulation. Additionally, circHMCU suppressed tumor growth in vivo.ConclusionCircHMCU acted as an oncogenic factor by regulating the miR-4458/PGK1 axis in BC.

Glycolysis-enhancing α1-adrenergic antagonists modify cognitive symptoms related to Parkinson’s disease

Terazosin is an α1-adrenergic receptor antagonist that enhances glycolysis and increases cellular ATP by binding to the enzyme phosphoglycerate kinase 1 (PGK1). Recent work has shown that terazosin is protective against motor dysfunction in rodent models of Parkinson’s disease (PD) and is associated with slowed motor symptom progression in PD patients. However, PD is also characterized by profound cognitive symptoms. We tested the hypothesis that terazosin protects against cognitive symptoms associated with PD. We report two main results. First, in rodents with ventral tegmental area (VTA) dopamine depletion modeling aspects of PD-related cognitive dysfunction, we found that terazosin preserved cognitive function. Second, we found that after matching for demographics, comorbidities, and disease duration, PD patients newly started on terazosin, alfuzosin, or doxazosin had a lower hazard of being diagnosed with dementia compared to tamsulosin, an α1-adrenergic receptor antagonist that does not enhance glycolysis. Together, these findings suggest that in addition to slowing motor symptom progression, glycolysis-enhancing drugs protect against cognitive symptoms of PD.

McPGK1-dependent mitochondrial import of PGK1 promotes metabolic reprogramming and self-renewal of liver TICs

Liver tumour-initiating cells (TICs) contribute to tumour initiation, metastasis, progression and drug resistance. Metabolic reprogramming is a cancer hallmark and plays vital roles in liver tumorigenesis. However, the role of metabolic reprogramming in TICs remains poorly explored. Here, we identify a mitochondria-encoded circular RNA, termed mcPGK1 (mitochondrial circRNA for translocating phosphoglycerate kinase 1), which is highly expressed in liver TICs. mcPGK1 knockdown impairs liver TIC self-renewal, whereas its overexpression drives liver TIC self-renewal. Mechanistically, mcPGK1 regulates metabolic reprogramming by inhibiting mitochondrial oxidative phosphorylation (OXPHOS) and promoting glycolysis. This alters the intracellular levels of α-ketoglutarate and lactate, which are modulators in Wnt/β-catenin activation and liver TIC self-renewal. In addition, mcPGK1 promotes PGK1 mitochondrial import via TOM40 interactions, reprogramming metabolism from oxidative phosphorylation to glycolysis through PGK1-PDK1-PDH axis. Our work suggests that mitochondria-encoded circRNAs represent an additional regulatory layer controlling mitochondrial function, metabolic reprogramming and liver TIC self-renewal.

PGK1 modulates balance between pro- and anti-inflammatory cytokines by interacting with ITI-H4

Inter-α-trypsin inhibitor heavy chain 4 (ITI-H4) is one of the acute phase proteins and is mainly related with inflammatory diseases such as bacterial bloodstream infection and recurrent pregnancy loss (RPL). In a previous study, ITI-H4 was reported to be cleaved by kallikrein B1 (KLKB1) and its cleaved form induces the imbalance between pro- and anti-inflammatory cytokines. Therefore, in this study, putative substrates of ITI-H4 were isolated by immunoprecipitation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) analysis. Of those, phosphoglycerate kinase 1 (PGK1) was found to be a binding protein of ITI-H4. PGK1 increases the level of ITI-H4 expression and blocks the cleavage of ITI-H4 mediated by KLKB1. It also inhibits pro-inflammatory response by inhibiting the JAK2/STAT3 signaling pathway. Therefore, PGK1, a novel binding partner of ITI-H4, is expected to have cellular functions in the pathogenesis of ITI-H4-related inflammatory diseases.