Phosphoglycerate Kinase Research Articles

Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins

ABSTRACT Macroautophagy/autophagy is a finely-regulated process in which cytoplasm encapsulated within transient organelles termed autophagosomes is delivered to lysosomes or vacuoles for degradation. Phospholipids, particularly phosphatidic acid (PA) that functions as a second messenger, play crucial and differential roles in autophagosome formation; however, the underlying mechanism remains largely unknown. Here we demonstrated that PA inhibits autophagy through competitive inhibition of the formation of ATG3 (autophagy-related)-ATG8e and ATG6-VPS34 (vacuolar protein sorting 34) complexes. PA bound to GAPC (glyceraldehyde-3-phosphate dehydrogenase) or PGK (phosphoglycerate kinase) and promoted their interaction with ATG3 or ATG6, which further attenuated the interactions of ATG3-ATG8e or ATG6-VPS34, respectively. Structural and mutational analyses revealed the mechanism of PA binding with GAPCs and PGK3, and that GAPCs or ATG8e competitively interacted with ATG3, and PGK3 or VPS34 competitively interacted with ATG6, at the same binding interface. These results elucidate the molecular mechanism of how PA inhibits autophagy through binding GAPC or PGK3 proteins and expand the understanding of the functional mode of PA, demonstrating the importance of phospholipids in plant autophagy and providing a new perspective for autophagy regulation by phospholipids. Abbreviation: ATG: autophagy-related; BiFC: bimolecular fluorescence complementation; co-IP: co-immunoprecipitation; Con A: concanamycin A; ER: endoplasmic reticulum; EZ: elongation zone; FRET-FLIM: fluorescence resonance energy transfer with fluorescence lifetime imaging microscopy; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GST: glutathione S-transferase; MDC: monodansylcadaverine; MZ: meristem zone; PA: phosphatidic acid; PAS: phagophore assembly site; PC: phosphatidylcholine; PE: phosphatidylethanolamine; PGK3: phosphoglycerate kinase; PtdIns3K: phosphatidylinositol 3-kinase; PLD: phospholipase D; TEM: transmission electron microscopy; TOR: target of rapamycin; VPS34: vacuolar protein sorting 34; WT: wild type; Y2H: yeast two-hybrid.

Circ_0076305 facilitates prostate cancer development via sponging miR-411-5p and regulating PGK1.

Abnormal expression of circular RNA (circRNA) is tightly linked to cancer progression. In this study, we aimed to investigate the biological role of circ_0076305 in prostate cancer (PCa). RT-qPCR was utilized to examine circ_0076305, microRNA-411-5p (miR-411-5p) and phosphoglycerate kinase 1 (PGK1) expression in PCa tissues and cells. CCK-8 assay, EdU assay, wound-healing assay and flow cytometry were executed to investigate the regulatory function of circ_0076305 on the proliferation, migration and apoptosis of PCa cells. Western blot (WB) assay was applied for measuring the protein levels. The effect of circ_0076305 on cellular glycolysis was examined using commercial kits. RNA immunoprecipitation (RIP) and dual-luciferase reporter assays were conducted for confirming the association between miR-411-5p and circ_0076305 or PGK1. The role of circ_0076305 in vivo was detected via establishing mice xenograft model. Circ_0076305 was highly expressed in PCa. Circ_0076305silencing could repress cell growth, migration and glycolysis while triggered apoptosis in PCa cells. MiR-411-5p was targeted by circ_0076305, and miR-411-5p suppression counteracted the influence of circ_0076305silencing in PCa cells. Additionally, miR-411-5p directly targeted PGK1, and miR-411-5p upregulation restrained PCa cell malignant behaviours via reducing PGK1. Mechanically, circ_0076305sponged miR-411-5p to affect PGK1 expression. Importantly, circ_0076305 interference inhibited tumour growth in vivo. Circ_0076305served as a novel oncogene PCa progression through regulation of miR-411-5p/PGK1 axis.

Disruption of Colorectal Cancer Network by Polyphyllins Reveals Pivotal Entities with Implications for Chemoimmunotherapy.

The prevalence of colorectal cancer has increased world-wide with high rates of mortality and morbidity. In the absence of efficacious drugs to treat this neoplasia, there is an imminent need to discover molecules with multifaceted effects. To this end, we opted to study the effect of steroidal saponins such as Polyphyllins. We performed anticancer activity studies with three analogs of Polyphyllins: Polyphyllin D (PD), Polyphyllin II (PII) and Polyphyllin G (PG). Here we show the potent effect of PD, PII (IC50 of 0.5−1 µM) and PG (IC50 of 3 µM) in inhibiting the viability of colorectal adenocarcinoma cells (DLD-1) and colorectal carcinoma cells (HCT116). PD and PII also showed inhibition of cell proliferation and sustained response upon withdrawal of the compounds when assessed by clonogenic assays in both the cell lines. Elucidation of the molecular mode of action revealed impact on the programmed cell death pathway. Additionally, proteomic profiling of DLD-1 revealed pivotal proteins differentially regulated by PD and PII, including a downregulated peroxiredoxin-1 which is considered as one of the novel targets to combat colorectal cancers and an upregulated elongation factor 2 (EF2), one of the key molecules considered as a tumor associated antigen (TAA) in colon cancer. Entities of cell metabolic pathways including downregulation of the key enzyme Phosphoglycerate kinase 1 of the glycolytic pathway was also observed. Importantly, the fold changes per se of the key components has led to the loss of viability of the colorectal cancer cells. We envision that the multifaceted function of PD and PII against the proliferation of colorectal carcinoma cells could have potential for novel treatments such as chemoimmunotherapy for colorectal adenocarcinomas. Future studies to develop these compounds as potent anti-colorectal cancer agents are warranted.

Jumonji-C domain-containing protein 5 suppresses proliferation and aerobic glycolysis in pancreatic cancer cells in a c-Myc-dependent manner

Despite the importance of metabolic reprogramming in cancer cells, the molecular mechanism regulating the tumor metabolic shift is still poorly understood. Deregulation of Jumonji-C domain-containing protein 5 (JMJD5) has been associated with multiple facets of biological processes in cancer cells. However, the role of JMJD5 in pancreatic cancer cells has seldom been discussed and requires further investigation. In the present study, by silencing or overexpressing JMJD5 in pancreatic cancer cells, we examined the impact of JMJD5 on cell proliferation and glucose metabolism. Using a dual luciferase assay, we assessed the effect of JMJD5 on the transcriptional activity of the c-Myc target gene. Analyzing The Cancer Genome Atlas and the Gene Expression Omnibus datasets revealed that low JMJD5 expression was associated with poor prognosis in patients with pancreatic cancer. JMJD5 loss promoted pancreatic cancer cell proliferation and induced a cellular metabolic shift from oxidative phosphorylation to glycolysis. In addition, in vivo experiments confirmed that ectopic JMJD5 expression inhibited cancer cell growth and the expression of glycolytic enzymes, such as lactate dehydrogenase and phosphoglycerate kinase 1. Moreover, JMJD5 negatively regulated c-Myc expression, the main regulator of cancer metabolism, leading to decreased c-Myc-targeted gene expression. Overall, the present study indicated that decreased JMJD5 expression promoted cell proliferation and glycolytic metabolism in pancreatic cancer cells in a c-Myc-dependent manner.

Zika Virus Infection of Sertoli Cells Alters Protein Expression Involved in Activated Immune and Antiviral Response Pathways, Carbohydrate Metabolism and Cardiovascular Disease.

Zika virus (ZIKV), a re-emerging virus, causes congenital brain abnormalities and Guillain–Barré syndrome. It is mainly transmitted by Aedes mosquitoes, but infections are also linked to sexual transmissions. Infectious ZIKV has been isolated, and viral RNA has been detected in semen over a year after the onset of initial symptoms, but the mode of long-term persistence is not yet understood. ZIKV can proliferate in human Sertoli cells (HSerC) for several weeks in vitro, suggesting that it might be a reservoir for persistent ZIKV infection. This study determined proteomic changes in HSerC during ZIKV infections by TMT-mass spectrometry analysis. Levels of 4416 unique Sertoli cell proteins were significantly altered at 3, 5, and 7 days after ZIKV infection. The significantly altered proteins include enzymes, transcription regulators, transporters, kinases, peptidases, transmembrane receptors, cytokines, ion channels, and growth factors. Many of these proteins are involved in pathways associated with antiviral response, antigen presentation, and immune cell activation. Several immune response pathway proteins were significantly activated during infection, e.g., interferon signaling, T cell receptor signaling, IL-8 signaling, and Th1 signaling. The altered protein levels were linked to predicted activation of immune response in HSerC, which was predicted to suppress ZIKV infection. ZIKV infection also affected the levels of critical regulators of gluconeogenesis and glycolysis pathways such as phosphoglycerate mutase, phosphoglycerate kinase, and enolase. Interestingly, many significantly altered proteins were associated with cardiac hypertrophy, which may induce heart failure in infected patients. In summary, our research contributes to a better understanding of ZIKV replication dynamics and infection in Sertoli cells.

PGK1 contributes to tumorigenesis and sorafenib resistance of renal clear cell carcinoma via activating CXCR4/ERK signaling pathway and accelerating glycolysis

Phosphoglycerate kinase 1 (PGK1) has complicated and multiple functions in cancer occurrence, tumor progression and drug resistance. Sorafenib is the first-line treatment targeted drug for patients with kidney renal clear cell carcinoma (KIRC) as a tyrosine kinase inhibitor, but sorafenib resistance is extremely common to retard therapy efficiency. So far, it is unclear whether and how PGK1 is involved in the pathogenesis and sorafenib resistance of KIRC. Herein, the molecular mechanisms of PGK1-mediated KIRC progression and sorafenib resistance have been explored by comprehensively integrative studies using biochemical approaches, mass spectrometry (MS) identification, microarray assay, nude mouse xenograft model and bioinformatics analysis. We have confirmed PGK1 is specifically upregulated in KIRC based on the transcriptome data generated by our own gene chip experiment, proteomics identification and the bioinformatics analysis for five online transcriptome datasets, and PGK1 upregulation in tumor tissues and serum is indicative with poor prognosis of KIRC patients. In the KIRC tissues, a high expression of PGK1 is often accompanied with an increase of glycolysis-related enzymes and CXCR4. PGK1 exhibits pro-tumorigenic properties in vitro and in a xenograft tumor model by accelerating glycolysis and inducing CXCR4-mediated phosphorylation of AKT and ERK. Moreover, PGK1 promotes sorafenib resistance via increasing CXCR4-mediated ERK phosphorylation. In conclusion, PGK1-invovled metabolic reprogramming and activation of CXCR4/ERK signaling pathway contributes to tumor growth and sorafenib resistance of KIRC.

A glycolytic enzyme complex and creatine kinase locally and differentially regulate ATP-sensitive potassium (KATP) channels in pancreatic islet alpha and beta cells

The ATP-sensitive potassium channel (KATP) is a key regulator of membrane potential and islet hormone secretion. Here, using the inside-out excised patch configuration, we isolated the plasma membrane compartment containing KATP on β-cells and α-cells from humans and mice. The characteristic response of KATP to inhibition by ATP and activation by ADP was used to identify the presence of a glycolytic enzyme complex that is functionally coupled to KATP regulation. We found that the phosphofructokinase product fructose-1,6-bisphosphate (FBP), which oscillates in β-cells, can induce KATP closure by its metabolism through the glycolytic chain including aldolase, GAPDH, phosphoglycerate kinase, phosphoglycerate mutase, enolase and finally pyruvate kinase.

Protein adaptation in extremophiles: a story of sequence, structure and osmolytes

Extremophiles are organisms that have optimal growth conditions in extreme temperature, pressure, acidity, salinity, or pH generally considered hostile for carbon-based life forms. Proteins have evolved to function in environments native to these different organisms. To understand how extremophiles have evolved their proteins we chose phosphoglycerate kinase (PGK), an essential enzyme in the glycolytic cycle from 6 different organisms. These were chosen to optimize for a variety of conditions across species, temperature and pressure optimal for their growth.

Aged Mouse Hippocampus Exhibits Signs of Chronic Hypoxia and an Impaired HIF-Controlled Response to Acute Hypoxic Exposures.

Altered hypoxia-inducible factor-alpha (HIF-α) activity may have significant consequences in the hippocampus, which mediates declarative memory, has limited vascularization, and is vulnerable to hypoxic insults. Previous studies have reported that neurovascular coupling is reduced in aged brains and that diseases which cause hypoxia increase with age, which may render the hippocampus susceptible to acute hypoxia. Most studies have investigated the actions of HIF-α in aging cortical structures, but few have focused on the role of HIF-α within aged hippocampus. This study tests the hypothesis that aging is associated with impaired hippocampal HIF-α activity. Dorsal hippocampal sections from mice aged 3, 9, 18, and 24 months were probed for the presence of HIF-α isoforms or their associated gene products using immunohistochemistry and fluorescent in situ hybridization (fISH). A subset of each age was exposed to acute hypoxia (8% oxygen) for 3 h to investigate changes in the responsiveness of HIF-α to hypoxia. Basal mean intensity of fluorescently labeled HIF-1α protein increases with age in the hippocampus, whereas HIF-2α intensity only increases in the 24-month group. Acute hypoxic elevation of HIF-1α is lost with aging and is reversed in the 24-month group. fISH reveals that glycolytic genes induced by HIF-1α (lactose dehydrogenase-a, phosphoglycerate kinase 1, and pyruvate dehydrogenase kinase 1) are lower in aged hippocampus than in 3-month hippocampus, and mRNA for monocarboxylate transporter 1, a lactose transporter, increases. These results indicate that lactate, used in neurotransmission, may be limited in aged hippocampus, concurrent with impaired HIF-α response to hypoxic events. Therefore, impaired HIF-α may contribute to age-associated cognitive decline during hypoxic events.

Proteome basis for the biological variations in color and tenderness of longissimus thoracis muscle from beef cattle differing in growth rate and feeding regime

The proteome basis for the biological variations in color and tenderness of longissimus thoracis muscle from ½ Angus (Bos taurus taurus) × ½ Nellore (Bos taurus indicus) crossbred steers was evaluated in a completely randomized experimental design consisting of four treatments (n = 9 per treatment): 1) feedlot finished, high growth rate (FH); 2) feedlot finished, low growth rate (FL); 3) pasture finished, high growth rate (PH); and 4) pasture finished, low growth rate (PL). The following comparisons were made to evaluate the effects of finishing systems and growth rates on muscle proteome: 1) FH × PL; 2) FL × PH; 3) FH × FL; and 4) PH × PL. Sixteen protein spots were differentially abundant among these comparisons (P ≤ 0.05), which were distinguished in two major clusters, energy metabolism- and muscle structure-related proteins that impacted glycolysis, carbon metabolism, amino acid biosynthesis and muscle contraction pathways (FDR ≤ 0.05). For FH × PL comparison, triosephosphate isomerase (TPI), phosphoglucomutase-1 (PGM1) and phosphoglycerate kinase 1 (PGK1) were overabundant in FH beef whereas troponin T (TNNT3), α-actin (ACTA1) and myosin regulatory light chain 2 (MYLPF) were overabundant in PL beef. For the FL × PH comparison, PGM1, phosphoglycerate mutase 2 (PGAM2) and annexin 2 (ANXA2) were overabundant in PH beef. For the FH × FL comparison, AMP deaminase (AMPD1) and serum albumin (ALB) were overabundant in FH beef whereas glycogen phosphorylase (PYGM) was overabundant in FL beef. For the PH × PL comparison, myoglobin (MB) was overabundant in PH beef whereas PYGM and MYLPF were overabundant in PL beef. In non-aged beef, L* was positively correlated with PGM1 (r = 0.54) while tenderness was negatively correlated with PGAM2 (r = -0.74) and ANXA2 (r = -0.60). In 7-d aged beef, color attributes (L*, a* and b*) were positively correlated with PGM1 (r = 0.67, 0.64 and 0.64, respectively) while tenderness was negatively correlated with TNNT3 (r = -0.57), PGK1 (r = -0.52) and MYLPF (r = -0.66). Therefore, finishing systems and growth rate affected the muscle proteome profile, which was related to beef color and tenderness. Additionally, these results suggest potential biomarkers for beef color (PGM1 and PGAM2) and tenderness (ANXA2, MYLPF, PGK1 and TNNT3).

LncRNA HCG18 promotes osteosarcoma growth by enhanced aerobic glycolysis via the miR-365a-3p/PGK1 axis

BackgroundOsteosarcoma (OS) is a common primary bone malignancy. Long noncoding RNA HCG18 is known to play an important role in a variety of cancers. However, its role in OS and relevant molecular mechanisms are unclear.MethodsReal-time quantitative PCR was performed to determine the expression of target genes. Function experiments showed the effects of HCG18 and miR-365a-3p on OS cell growth.ResultsHCG18 expression was increased in OS cell lines. Moreover, in vitro and in vivo experiments demonstrated that HCG18 knockdown inhibited OS cell proliferation. Mechanistically, HCG18 was defined as a competing endogenous RNA by sponging miR-365a-3p, thus elevating phosphoglycerate kinase 1 (PGK1) expression by directly targeting its 3ʹUTR to increase aerobic glycolysis.ConclusionHCG18 promoted OS cell proliferation via enhancing aerobic glycolysis by regulating the miR-365a-3p/PGK1 axis. Therefore, HCG18 may be a potential target for OS treatment.

Hereditary myopathies associated with hematological abnormalities.

The diagnostic evaluation of a patient with suspected hereditary muscle disease can be challenging. Clinicians rely largely on clinical history and examination features, with additional serological, electrodiagnostic, radiologic, histopathologic, and genetic investigations assisting in definitive diagnosis. Hematological testing is inexpensive and widely available, but frequently overlooked in the hereditary myopathy evaluation. Hematological abnormalities are infrequently encountered in this setting; however, their presence provides a valuable clue, helps refine the differential diagnosis, tailors further investigation, and assists interpretation of variants of uncertain significance. A diverse spectrum of hematological abnormalities is associated with hereditary myopathies, including anemias, leukocyte abnormalities, and thrombocytopenia. Recurrent rhabdomyolysis in certain glycolytic enzymopathies co-occurs with hemolytic anemia, often chronic and mild in phosphofructokinase and phosphoglycerate kinase deficiencies, or acute and fever-associated in aldolase-A and triosephosphate isomerase deficiency. Sideroblastic anemia, commonly severe, accompanies congenital-to-childhood onset mitochondrial myopathies including Pearson marrow-pancreas syndrome and mitochondrial myopathy, lactic acidosis, and sideroblastic anemia phenotypes. Congenital megaloblastic macrocytic anemia and mitochondrial dysfunction characterize SFXN4-related myopathy. Neutropenia, chronic or cyclical, with recurrent infections, infantile-to-childhood onset skeletal myopathy and cardiomyopathy are typical of Barth syndrome, while chronic neutropenia without infection occurs rarely in DNM2-centronuclear myopathy. Peripheral eosinophilia may accompany eosinophilic inflammation in recessive calpainopathy. Lipid accumulation in leukocytes on peripheral blood smear (Jordans' anomaly) is pathognomonic for neutral lipid storage diseases. Mild thrombocytopenia occurs in autosomal dominant, childhood-onset STIM1 tubular aggregate myopathy, STIM1 and ORAI1 deficiency syndromes, and GNE myopathy. Herein, we review these hereditary myopathies in which hematological features play a prominent role.

Comparative analysis of proteins associated with 26S and 20S proteasomes isolated from rabbit brain and liver

We have isolated fractions of 26S and 20S proteasomes were from the rabbit liver and the brain. According to mass spectrometric (MS) analysis, the 26S proteasome fractions from these organs contained catalytic and regulatory subunits characteristic of the proteasome core and regulatory subunits. The 20S fractions of brain and liver proteasomes contained only catalytic proteasome subunits. In addition to proteasome subunits, the isolated fractions contained components of the ubiquitin-proteasome system, ubiquitinated proteins, enzymes that play an important role in metabolic processes, cytoskeletal components, signaling, regulatory, and protective proteins, as well as proteins regulating gene expression, cell division, and differentiation. The abundance of a number of proteasome-associated proteins was comparable or exceeded the abundance of intrinsic proteasome components. About a third of the proteins common to all studied fractions (26S and 20S of brain and liver proteasomes) belong to the group of multifunctional proteins. Selective biosensor validation confirmed the affinity binding of proteins (aldolase, phosphoglycerate kinase) identified during MS analysis to the brain 20S proteasome. Comparison of the subproteomes of the 26S and 20S brain proteasomes showed that removal of components of the regulatory (19S) subparticles caused almost two-fold increase in the total number of individual proteins associated with the core part of the proteasome (20S). In the liver, the number of proteins associated with the core part of the proteasome remained basically unchanged after the removal of the components of the regulatory (19S) subparticles. This indicates that in the brain and, possibly, in other organs, proteins of the regulatory (19S) subunit play an important role in the formation of the proteasome interactome.

Comparative analysis of proteins associated with 26S and 20S proteasomes isolated from rabbit brain and liver

We have isolated fractions of 26S and 20S proteasomes were from the rabbit liver and the brain. According to mass spectrometric (MS) analysis, the 26S proteasome fractions from these organs contained catalytic and regulatory subunits characteristic of the proteasome core and regulatory subunits. The 20S fractions of brain and liver proteasomes contained only catalytic proteasome subunits. In addition to proteasome subunits, the isolated fractions contained components of the ubiquitin-proteasome system, ubiquitinated proteins, enzymes that play an important role in metabolic processes, cytoskeletal components, signaling, regulatory, and protective proteins, as well as proteins regulating gene expression, cell division, and differentiation. The abundance of a number of proteasome-associated proteins was comparable or exceeded the abundance of intrinsic proteasome components. About a third of the proteins common to all studied fractions (26S and 20S of brain and liver proteasomes) belong to the group of multifunctional proteins. Selective biosensor validation confirmed the affinity binding of proteins (aldolase, phosphoglycerate kinase) identified during MS analysis to the brain 20S proteasome. Comparison of the subproteomes of the 26S and 20S brain proteasomes showed that removal of components of the regulatory (19S) subparticles caused almost two-fold increase in the total number of individual proteins associated with the core part of the proteasome (20S). In the liver, the number of proteins associated with the core part of the proteasome remained basically unchanged after the removal of the components of the regulatory (19S) subparticles. This indicates that in the brain and, possibly, in other organs, proteins of the regulatory (19S) subunit play an important role in the formation of the proteasome interactome.Iz pecheni i mozga krolika vydeleny fraktsii 26S i 20S proteasom. Po dannym mass-spektrometricheskogo (MS) analiza, fraktsii 26S proteasom iz étikh organov soderzhali kataliticheskie i reguliatornye sub"edinitsy, kharakternye dlia korovoĭ chasti proteasom i dlia reguliatornykh subchastits. Fraktsii 20S proteasom mozga i pecheni soderzhali tol'ko kataliticheskie sub"edinitsy proteasom. Pomimo sub"edinits proteasom, v poluchennykh fraktsiiakh obnaruzheny komponenty ubikvitin-proteasomnoĭ sistemy, ubikvitinirovannye belki, fermenty, igraiushchie vazhnuiu rol' v metabolicheskikh protsessakh, komponenty tsitoskeleta, signal'nye, reguliatornye i zashchitnye belki, a takzhe belki reguliatsii ékspressii genov, kletochnogo deleniia i differentsirovki. Predstavlennost' riada assotsiirovannykh s proteasomami belkov byla sopostavima ili prevyshala predstavlennost' sobstvennykh komponentov proteasom. Okolo treti belkov, obshchikh dlia vsekh issledovannykh fraktsiĭ (26S i 20S proteasom mozga i pecheni), otnosiatsia k gruppe mul'tifunktsional'nykh belkov. Vyborochnaia biosensornaia validatsiia podtverdila affinnoe sviazyvanie identifitsirovannykh v khode MS analiza belkov (al'dolaza, fosfoglitseratkinaza) s 20S proteasomoĭ mozga. Sopostavlenie subproteomov 26S i 20S proteasom mozga pokazalo, chto pri udalenii komponentov reguliatornykh (19S) subchastits kolichestvo individual'nykh belkov, assotsiirovannykh s korovoĭ chast'iu proteasomy (20S), prakticheski udvaivalos'. V pecheni kolichestvo belkov, assotsiirovannykh s korovoĭ chast'iu proteasomy, posle udaleniia komponentov reguliatornykh subchastits prakticheski ne izmenialos'. Éto svidetel'stvuet o tom, chto v mozge i, vozmozhno, v drugikh organakh belki reguliatornoĭ (19S) subchastitsy igraiut vazhnuiu rol' v formirovanii proteasomnogo interaktoma.

Overexpression of glycolysis markers in placental tissue of pregnant women with chronic venous disease: a histological study.

Chronic Venous Disease (CVD) refers to a wide variety of venous disorders being the varicose veins its most common manifestation. It is well-established the link between pregnancy and the risk of suffering CVD, due to hormonal or haematological factors, especially during the third trimester. In the same manner, previous studies have demonstrated the detrimental effect of this condition in the placental tissue of pregnant women, including in the normal physiology and the metabolomic profile of this organ. In this context, the aim of this study was to evaluate the glucose homeostasis in the placental tissue of women presenting CVD. Through immunohistochemistry, we studied the protein expression of the glucose transporter 1 (GLUT-1), Phosphoglycerate kinase 1 (PGK1), aldolase (ALD), Glyceraldehyde-3-phosphate dehydrogenase (GA3PDH) and lactate dehydrogenase (LDH). Our results have reported a significative increase in the expression of GLUT-1, PGK1, ALD, GA3PDH and the isoenzyme LDHA in placentas of women with CVD. This work has proven for the first-time an altered glucose metabolism in the placental tissue of women affected by CVD, what may aid to understand the pathophysiological mechanisms of this condition in more distant organs such as placenta. Furthermore, our research also supports the basis for further studies in the metabolic phenotyping of the human placenta due to CVD, which may be considered during the late pregnancy in these women.

PGK1 represses autophagy-mediated cell death to promote the proliferation of liver cancer cells by phosphorylating PRAS40

Autophagy predominantly promotes cell survival by recycling cell components, while it kills cells in specific contexts. Cell death related to autophagy plays important roles in multiple physiological and pathological situations including tumorigenesis, and the mechanism needs to be defined further. PRAS40 was found to be crucial in various cancers, and phosphorylation was reported to be involved in autophagy inhibition in monocytes. However, the detailed role of PRAS40 in autophagy and the relationship to tumorigenesis remain largely unknown. Herein we screened the binding partners of PRAS40, and found that PRAS40 interacted with Phosphoglycerate kinase 1 (PGK1). PGK1 phosphorylated PRAS40 at Threonine 246, which could be inhibited by blocking the interaction. Both in vitro and in vivo results revealed that PRAS40 mediated PGK1-induced cell growth. By tracing the mechanism, we found that PGK1 suppressed autophagy-mediated cell death, in which PRAS40 was crucial. Thus PGK1 phosphorylates PRAS40 to repress autophagy-mediated cell death under normoxia, promoting cellular proliferation. The binding of PGK1 to PRAS40 was transferred to Beclin1 under hypoxia, resulting in the increase of Beclin1 phosphorylation. These results suggest a novel model of tumorigenesis, in which PGK1 switches between repressing autophagy-mediated cell death via PRAS40 and inducing autophagy through Beclin1 according to the environmental oxygen level. Our study is anticipated to be able to offer novel insights in understanding PGK1/PRAS40 signaling hyperactivated cancers.

Activating Silent Glycolysis Bypasses in Escherichia coli.

All living organisms share similar reactions within their central metabolism to provide precursors for all essential building blocks and reducing power. To identify whether alternative metabolic routes of glycolysis can operate in E. coli, we complementarily employed in silico design, rational engineering, and adaptive laboratory evolution. First, we used a genome-scale model and identified two potential pathways within the metabolic network of this organism replacing canonical Embden-Meyerhof-Parnas (EMP) glycolysis to convert phosphosugars into organic acids. One of these glycolytic routes proceeds via methylglyoxal and the other via serine biosynthesis and degradation. Then, we implemented both pathways in E. coli strains harboring defective EMP glycolysis. Surprisingly, the pathway via methylglyoxal seemed to immediately operate in a triosephosphate isomerase deletion strain cultivated on glycerol. By contrast, in a phosphoglycerate kinase deletion strain, the overexpression of methylglyoxal synthase was necessary to restore growth of the strain. Furthermore, we engineered the "serine shunt" which converts 3-phosphoglycerate via serine biosynthesis and degradation to pyruvate, bypassing an enolase deletion. Finally, to explore which of these alternatives would emerge by natural selection, we performed an adaptive laboratory evolution study using an enolase deletion strain. Our experiments suggest that the evolved mutants use the serine shunt. Our study reveals the flexible repurposing of metabolic pathways to create new metabolite links and rewire central metabolism.

HPV16 E6/E7 stabilize PGK1 protein by reducing its poly-ubiquitination in cervical cancer.

This study aimed to explore the expression profile, prognostic value, regulatory effect, and the underlying mechanism of dysregulation of phosphoglycerate kinase 1 (PGK1) in high-risk human papillomavirus (HPV)-positive cervical epithelial squamous cell carcinoma (CESC). Bioinformatic analysis was performed using the CESC subset of The Cancer Genome Atlas (TCGA)-Cervical Cancer (CESC) and normal cervix in The Genotype-Tissue Expression (GTEx) project. HPV-16 positive CaSki and SiHa cells were used as in vitro cell models. Results showed that compared to the normal cervix, CESC tissues had significantly higher expression of PGK1. CESC patients with the higher 50% expression of PGK1 had substantially shorter disease-specific survival (DSS), and progression-free survival (PFS) compared to the cases with the lower 50% expression of PGK1. PGK1 knockdown impaired, but PGK1 overexpression enhanced the proliferation, colony formation, aerobic glycolytic activities (lactate production, intracellular ATP levels, glucose uptake, and extracellular acidification rate), migration, and invasion of CaSki and SiHa cells. HPV-16 E6/E7 knockdown in CaSki and SiHa cells had limited influence on PGK1 transcription but significantly decreased the half-life of PGK1 protein. E6/E7 knockdown mediated PGK1 downregulation could be blocked by adding MG-132. PGK1 poly-ubiquitination was significantly enhanced after E6/E7 knockdown. In conclusion, this study showed that PGK1 expression might serve as a prognostic biomarker in cervical cancer. Its upregulation contributes to enhanced aerobic glycolysis, migration, and invasion of CESC cells. HPV16 E6/E7 stabilizes PGK1 protein by reducing its poly-ubiquitination.

Molecular mechanism of glycolytic flux control intrinsic to human phosphoglycerate kinase

Glycolysis plays a fundamental role in energy production and metabolic homeostasis. The intracellular [adenosine triphosphate]/[adenosine diphosphate] ([ATP]/[ADP]) ratio controls glycolytic flux; however, the regulatory mechanism underlying reactions catalyzed by individual glycolytic enzymes enabling flux adaptation remains incompletely understood. Phosphoglycerate kinase (PGK) catalyzes the reversible phosphotransfer reaction, which directly produces ATP in a near-equilibrium step of glycolysis. Despite extensive studies on the transcriptional regulation of PGK expression, the mechanism in response to changes in the [ATP]/[ADP] ratio remains obscure. Here, we report a protein-level regulation of human PGK (hPGK) by utilizing the switching ligand-binding cooperativities between adenine nucleotides and 3-phosphoglycerate (3PG). This was revealed by nuclear magnetic resonance (NMR) spectroscopy at physiological salt concentrations. MgADP and 3PG bind to hPGK with negative cooperativity, whereas MgAMPPNP (a nonhydrolyzable ATP analog) and 3PG bind to hPGK with positive cooperativity. These opposite cooperativities enable a shift between different ligand-bound states depending on the intracellular [ATP]/[ADP] ratio. Based on these findings, we present an atomic-scale description of the reaction scheme for hPGK under physiological conditions. Our results indicate that hPGK intrinsically modulates its function via ligand-binding cooperativities that are finely tuned to respond to changes in the [ATP]/[ADP] ratio. The alteration of ligand-binding cooperativities could be one of the self-regulatory mechanisms for enzymes in bidirectional pathways, which enables rapid adaptation to changes in the intracellular environment.

Anaesthesia Challenges for a Patient with Phosphoglycerate Kinase Deficiency Undergoing Open Gastrostomy Procedure: A Case Report.

Phosphoglycerate kinase deficiency (PGK1D) is a rare X-linked metabolic disorder with variable phenotype. Mutations of the PGK1 gene result in clinically variable spherocytic hemolytic anemia and variable defects in the central nervous system. Rhabdomyolysis, myopathy, migraine, and retinal involvement have also been reported as clinical consequences. We describe for the first time the anaesthetic management of a patient with the X-linked phosphoglycerate kinase deficiency scheduled for an open gastrostomy procedure to facilitate enteral nutrition due to chronic oral aversion. Given the various organs systems involved, we suggest a number of preoperative investigations and describe our intraoperative management. Given the paucity of literature regarding children with this condition, we believe this case report will be a worthwhile addition to the anaesthetic literature to help guide other anaesthetists managing patients with this condition.