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Neuro-oncology Feb 2023
Topics: Humans; Bevacizumab; Phosphofructokinase-1; Phosphorylation; Neoplasms
PubMed: 36516222
DOI: 10.1093/neuonc/noac269 -
Advances in Biological Regulation Dec 2023Phosphofructokinase is the central enzyme in glycolysis and constitutes a highly regulated step. The liver isoform (PFKL) compartmentalizes during activation and...
Phosphofructokinase is the central enzyme in glycolysis and constitutes a highly regulated step. The liver isoform (PFKL) compartmentalizes during activation and inhibition in vitro and in vivo, respectively. Compartmentalized PFKL is hypothesized to modulate metabolic flux consistent with its central role as the rate limiting step in glycolysis. PFKL tetramers self-assemble at two interfaces in the monomer (interface 1 and 2), yet how these interfaces contribute to PFKL compartmentalization and drive protein interactions remains unclear. Here, we used site-specific incorporation of noncanonical photocrosslinking amino acids to identify PFKL interactors at interface 1, 2, and the active site. Tandem mass tag-based quantitative interactomics reveals interface 2 as a hotspot for PFKL interactions, particularly with cytoskeletal, glycolytic, and carbohydrate derivative metabolic proteins. Furthermore, PFKL compartmentalization into puncta was observed in human cells using citrate inhibition. Puncta formation attenuated crosslinked protein-protein interactions with the cytoskeleton at interface 2. This result suggests that PFKL compartmentalization sequesters interface 2, but not interface 1, and may modulate associated protein assemblies with the cytoskeleton.
Topics: Humans; Phosphofructokinases; Phosphofructokinase-1; Liver; Citrates; Citric Acid
PubMed: 37806136
DOI: 10.1016/j.jbior.2023.100987 -
FASEB Journal : Official Publication of... Oct 2023A link between increased glycolysis and vascular calcification has recently been reported, but it remains unclear how increased glycolysis contributes to vascular...
A link between increased glycolysis and vascular calcification has recently been reported, but it remains unclear how increased glycolysis contributes to vascular calcification. We therefore investigated the role of PFKFB3, a critical enzyme of glycolysis, in vascular calcification. We found that PFKFB3 expression was upregulated in calcified mouse VSMCs and arteries. We showed that expression of miR-26a-5p and miR-26b-5p in calcified mouse arteries was significantly decreased, and a negative correlation between Pfkfb3 mRNA expression and miR-26a-5p or miR-26b-5p was seen in these samples. Overexpression of miR-26a/b-5p significantly inhibited PFKFB3 expression in VSMCs. Intriguingly, pharmacological inhibition of PFKFB3 using PFK15 or knockdown of PFKFB3 ameliorated vascular calcification in vD -overloaded mice in vivo or attenuated high phosphate (Pi)-induced VSMC calcification in vitro. Consistently, knockdown of PFKFB3 significantly reduced glycolysis and osteogenic transdifferentiation of VSMCs, whereas overexpression of PFKFB3 in VSMCs induced the opposite effects. RNA-seq analysis and subsequent experiments revealed that silencing of PFKFB3 inhibited FoxO3 expression in VSMCs. Silencing of FoxO3 phenocopied the effects of PFKFB3 depletion on Ocn and Opg expression but not Alpl in VSMCs. Pyruvate or lactate supplementation, the product of glycolysis, reversed the PFKFB3 depletion-mediated effects on ALP activity and OPG protein expression in VSMCs. Our results reveal that blockade of PFKFB3-mediated glycolysis inhibits vascular calcification in vitro and in vivo. Mechanistically, we show that FoxO3 and lactate production are involved in PFKFB3-driven osteogenic transdifferentiation of VSMCs. PFKFB3 may be a promising therapeutic target for the treatment of vascular calcification.
Topics: Animals; Mice; Glycolysis; Lactic Acid; MicroRNAs; Muscle, Smooth, Vascular; Phosphoric Monoester Hydrolases; Vascular Calcification; Phosphofructokinase-2; Forkhead Box Protein O3
PubMed: 37682013
DOI: 10.1096/fj.202300900R -
Frontiers in Immunology 2022Alcohol abuse, reported by 1/8 critically ill patients, is an independent risk factor for death in sepsis. Sepsis kills over 270,000 patients/year in the US. We reported...
Alcohol abuse, reported by 1/8 critically ill patients, is an independent risk factor for death in sepsis. Sepsis kills over 270,000 patients/year in the US. We reported that the ethanol-exposure suppresses innate-immune response, pathogen clearance, and decreases survival in sepsis-mice sirtuin 2 (SIRT2). SIRT2 is an NAD+-dependent histone-deacetylase with anti-inflammatory properties. We hypothesized that in ethanol-exposed macrophages, SIRT2 suppresses phagocytosis and pathogen clearance by regulating glycolysis. Immune cells use glycolysis to fuel increased metabolic and energy demand of phagocytosis. Using ethanol-exposed mouse bone marrow- and human blood monocyte-derived macrophages, we found that SIRT2 mutes glycolysis deacetylating key glycolysis regulating enzyme phosphofructokinase-platelet isoform (PFKP), at mouse lysine 394 (mK394, human: hK395). Acetylation of PFKP at mK394 (hK395) is crucial for PFKP function as a glycolysis regulating enzyme. The PFKP also facilitates phosphorylation and activation of autophagy related protein 4B (Atg4B). Atg4B activates microtubule associated protein 1 light chain-3B (LC3). LC3 is a driver of a subset of phagocytosis, the LC3-associated phagocytosis (LAP), which is crucial for segregation and enhanced clearance of pathogens, in sepsis. We found that in ethanol-exposed cells, the SIRT2-PFKP interaction leads to decreased Atg4B-phosphorylation, decreased LC3 activation, repressed phagocytosis and LAP. Genetic deficiency or pharmacological inhibition of SIRT2 reverse PFKP-deacetylation, suppressed LC3-activation and phagocytosis including LAP, in ethanol-exposed macrophages to improve bacterial clearance and survival in ethanol with sepsis mice.
Topics: Animals; Humans; Mice; Ethanol; Macrophages; Phagocytosis; Sirtuin 2; Phosphofructokinase-1, Type C
PubMed: 36865524
DOI: 10.3389/fimmu.2022.1079962 -
American Journal of Physiology. Cell... Nov 2023Glomerular angiogenesis is a characteristic feature of diabetic nephropathy (DN). Enhanced glycolysis plays a crucial role in angiogenesis. The present study was...
Glomerular angiogenesis is a characteristic feature of diabetic nephropathy (DN). Enhanced glycolysis plays a crucial role in angiogenesis. The present study was designed to investigate the role of glycolysis in glomerular endothelial cells (GECs) in a mouse model of DN. Mouse renal cortex and isolated glomerular cells were collected for single-cell and RNA sequencing. Cultured GECs were exposed to high glucose in the presence (proangiogenic) and absence of a vascular sprouting regimen. MicroRNA-590-3p was delivered by lipofectamine in vivo and in vitro. In the present study, a subgroup of GECs with proangiogenic features was identified in diabetic kidneys by using sequencing analyses. In cultured proangiogenic GECs, high glucose increased glycolysis and phosphofructokinase/fructose bisphosphatase 3 (PFKFB3) protein expression, which were inhibited by overexpressing miRNA-590-3p. Mimics of miRNA-590-3p also increased receptor for sphingosine 1-phosphate (S1pR1) expression, an angiogenesis regulator, in proangiogenic GECs challenged with high glucose. Inhibition of PFKFB3 by pharmacological and genetic approaches upregulated S1pR1 protein in vitro. Mimics of miRNA-590-3p significantly reduced migration and angiogenic potential in proangiogenic GECs challenged with high glucose. Ten-week-old type 2 diabetic mice had elevated urinary albumin levels, reduced renal cortex miRNA-590-3p expression, and disarrangement of glomerular endothelial cell fenestration. Overexpressing miRNA-590-3p via perirenal adipose tissue injection restored endothelial cell fenestration and reduced urinary albumin levels in diabetic mice. Therefore, the present study identifies a subgroup of GECs with proangiogenic features in mice with DN. Local administration of miRNA-590-3p mimics reduces glycolytic rate and upregulates S1pR1 protein expression in proangiogenic GECs. The protective effects of miRNA-590-3p provide therapeutic potential in DN treatment. Proangiogenetic glomerular endothelial cells (GECs) are activated in diabetic nephropathy. High glucose upregulates glycolytic enzyme phosphofructokinase/fructose bisphosphatase 3 (PFKFB3) in proangiogenetic cells. PFKFB3 protects the glomerular filtration barrier by targeting endothelial S1pR1. MiRNA-590-3p restores endothelial cell function and mitigates diabetic nephropathy.
Topics: Mice; Animals; Diabetic Nephropathies; Endothelial Cells; Fructose-Bisphosphatase; Phosphofructokinases; Diabetes Mellitus, Experimental; Sphingosine-1-Phosphate Receptors; Phosphofructokinase-1; Glucose; MicroRNAs; Albumins; Glycolysis
PubMed: 37781737
DOI: 10.1152/ajpcell.00261.2023 -
Cancer Metastasis Reviews Jun 2022Reprogrammed metabolism and high energy demand are well-established properties of cancer cells that enable tumor growth. Glycolysis is a primary metabolic pathway that... (Review)
Review
Reprogrammed metabolism and high energy demand are well-established properties of cancer cells that enable tumor growth. Glycolysis is a primary metabolic pathway that supplies this increased energy demand, leading to a high rate of glycolytic flux and a greater dependence on glucose in tumor cells. Finding safe and effective means to control glycolytic flux and curb cancer cell proliferation has gained increasing interest in recent years. A critical step in glycolysis is controlled by the enzyme 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), which converts fructose 6-phosphate (F6P) to fructose 2,6-bisphosphate (F2,6BP). F2,6BP allosterically activates the rate-limiting step of glycolysis catalyzed by PFK1 enzyme. PFKFB3 is often overexpressed in many human cancers including pancreatic, colon, prostate, and breast cancer. Hence, PFKFB3 has gained increased interest as a compelling therapeutic target. In this review, we summarize and discuss the current knowledge of PFKFB3 functions, its role in cellular pathways and cancer development, its transcriptional and post-translational activity regulation, and the multiple pharmacologic inhibitors that have been used to block PFKFB3 activity in cancer cells. While much remains to be learned, PFKFB3 continues to hold great promise as an important therapeutic target either as a single agent or in combination with current interventions for breast and other cancers.
Topics: Breast Neoplasms; Fructose; Glucose; Glycolysis; Humans; Male; Phosphofructokinase-2
PubMed: 35419769
DOI: 10.1007/s10555-022-10027-5 -
The Journal of Physical Chemistry. B Jun 2021Temporal order in living matters reflects the self-organizing nature of dynamical processes driven out of thermodynamic equilibrium. Because of functional reasons, the...
Temporal order in living matters reflects the self-organizing nature of dynamical processes driven out of thermodynamic equilibrium. Because of functional reasons, the period of a biochemical oscillation must be tuned to a specific value with precision; however, according to the thermodynamic uncertainty relation (TUR), the precision of the oscillatory period is constrained by the thermodynamic cost of generating it. After reviewing the basics of chemical oscillations using the Brusselator as a model system, we study the glycolytic oscillation generated by octameric phosphofructokinase (PFK), which is known to display a period of several minutes. By exploring the phase space of glycolytic oscillations, we find that the glycolytic oscillation under the cellular condition is realized in a cost-effective manner. Specifically, over the biologically relevant range of parameter values of glycolysis and octameric PFK, the entropy production from the glycolytic oscillation is minimal when the oscillation period is (5-10) min. Furthermore, the glycolytic oscillation is found at work near the phase boundary of limit cycles, suggesting that a moderate increase of glucose injection rate leads to the loss of oscillatory dynamics, which is reminiscent of the loss of pulsatile insulin release resulting from elevated blood glucose level.
Topics: Glycolysis; Models, Biological; Phosphofructokinase-1; Phosphofructokinases; Thermodynamics
PubMed: 34038120
DOI: 10.1021/acs.jpcb.1c01325 -
Cell Death & Disease Sep 2022Endometriosis (EM) is one of the vanquished wonted causes of chronic pelvic sting in women and is closely associated with infertility. The long-term, complex, systemic,...
Endometriosis (EM) is one of the vanquished wonted causes of chronic pelvic sting in women and is closely associated with infertility. The long-term, complex, systemic, and post-treatment recurrence of EM wreaks havoc on women's quality of life. Extensive metabolic reprogramming (aerobic glycolysis, glucose overweening intake, and high lactate production) and cancer-like changes have been found in EM, which bears striking similarities to tumorigenesis. The key glycolysis regulator PFKFB4 is overexpressed in EM. However, the mechanism of PFKFB4 in EM remains unknown. We found that PFKFB4 was upregulated and was closely related to the progression of EM. We identified focus PIM2 as a new pioneering adjoin protein of PFKFB4. Vigorous biochemical methods were used to confirm that PIM2 phosphorylated site Thr140 of PFKFB4. PIM2 also could enhance PFKFB4 protein expression through the ubiquitin-proteasome pathway. Moreover, PIM2 expression was really corresponding prevalent with PFKFB4 in endometriosis in vivo. Importantly, phosphorylation of PFKFB4 on Thr140 by PIM2 promoted EM glycolysis and cell growth. Our study demonstrates that PIM2 mediates PFKFB4 Thr140 phosphorylation thus regulating glycolysis and EM progression. We illustrated a new mechanism that PIM2 simulated a central upstream partnership in the regulation of PFKFB4, and reveal a novel means of PIM2-PFKFB4 setting EM growth. Our research provided new theoretical support for further clarifying the reprogramming of EM glucose metabolism, and provided new clues for exploring non-contraceptive treatments for EM.
Topics: Anaerobiosis; Cell Proliferation; Endometriosis; Female; Glucose; Glycolysis; Humans; Lactates; Phosphofructokinase-2; Phosphoric Monoester Hydrolases; Phosphorylation; Proteasome Endopeptidase Complex; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Quality of Life; Ubiquitins
PubMed: 36109523
DOI: 10.1038/s41419-022-05241-6 -
JCI Insight Dec 2022Acute lung injury (ALI) is a severe form of lung inflammation causing acute respiratory distress syndrome in patients. ALI pathogenesis is closely linked to uncontrolled...
Acute lung injury (ALI) is a severe form of lung inflammation causing acute respiratory distress syndrome in patients. ALI pathogenesis is closely linked to uncontrolled alveolar inflammation. We hypothesize that specific enzymes of the glycolytic pathway could function as key regulators of alveolar inflammation. Therefore, we screened isolated alveolar epithelia from mice exposed to ALI induced by injurious ventilation to assess their metabolic responses. These studies pointed us toward a selective role for isoform 3 of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3). Pharmacologic inhibition or genetic deletion of Pfkfb3 in alveolar epithelia (Pfkfb3loxP/loxP SPC-ER-Cre+ mice) was associated with profound increases in ALI during injurious mechanical ventilation or acid instillation. Studies in genetic models linked Pfkfb3 expression and function to Hif1a. Not only did intratracheal pyruvate instillation reconstitute Pfkfb3loxP/loxP or Hif1aloxP/loxP SPC-ER-Cre+ mice, but pyruvate was also effective in ALI treatment of wild-type mice. Finally, proof-of-principle studies in human lung biopsies demonstrated increased PFKFB3 staining in injured lungs and colocalized PFKFB3 to alveolar epithelia. These studies reveal a specific role for PFKFB3 in counterbalancing alveolar inflammation and lay the groundwork for novel metabolic therapeutic approaches during ALI.
Topics: Humans; Animals; Mice; Lung; Acute Lung Injury; Pneumonia; Inflammation; Phosphofructokinase-2; Hypoxia-Inducible Factor 1, alpha Subunit
PubMed: 36326834
DOI: 10.1172/jci.insight.157855 -
Frontiers in Bioscience (Landmark... Nov 2023The dilation of lymphatic vessels plays a critical role in maintaining heart function, while a lack thereof could contribute to heart failure (HF), and subsequently to...
BACKGROUND
The dilation of lymphatic vessels plays a critical role in maintaining heart function, while a lack thereof could contribute to heart failure (HF), and subsequently to an acute myocardial infarction (AMI). Macrophages participate in the induction of lymphangiogenesis by secreting vascular endothelial cell growth factor C (VEGF-C), although the precise mechanism remains unclear.
METHODS
Intramyocardial injections of adeno-associated viruses (AAV9) to inhibit the expression of ( shRNA) or promote the expression of ( ORF) in the heart; Myh6-mCherry B6 D2-tg mice and flow cytometry were used to evaluate the number of myocellular debris in the mediastinal lymph nodes; fluorescence staining and qPCR were used to evaluate fluorescence analysis; seahorse experiment was used to evaluate the level of glycolysis of macrophages; , , and mice were used as a model to knock out the expression of and in macrophages.
RESULTS
The escalation of in cardiac tissue can facilitate the drainage of myocardial debris to the mediastinal lymph nodes, thereby improving cardiac function and reducing fibrosis after reperfusion injury. Conversely, myeloid deficiency displayed an increase in macrophage counts and inflammation levels following reperfusion injury. The inhibition of the critical enzyme in macrophage glycolysis can stimulate the manifestation of in macrophages. A deficiency in myeloid is associated with induced lymphangiogenesis following reperfusion injury.
CONCLUSIONS
Our initial investigations suggest that the suppression of expression in macrophages could potentially stimulate the production of in these immune cells, which in turn may facilitate lymphangiogenesis and mitigate the inflammatory effects of I/R injury.
Topics: Animals; Mice; Lymphangiogenesis; Macrophages; Myocardial Infarction; Reperfusion Injury; Vascular Endothelial Growth Factor C; Phosphofructokinase-2
PubMed: 38062830
DOI: 10.31083/j.fbl2811277