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Nature Communications Jan 2017It is well known that c-Src has important roles in tumorigenesis. However, it remains unclear whether c-Src contributes to metabolic reprogramming. Here we find that...
It is well known that c-Src has important roles in tumorigenesis. However, it remains unclear whether c-Src contributes to metabolic reprogramming. Here we find that c-Src can interact with and phosphorylate hexokinases HK1 and HK2, the rate-limiting enzymes in glycolysis. Tyrosine phosphorylation dramatically increases their catalytic activity and thus enhances glycolysis. Mechanistically, c-Src phosphorylation of HK1 at Tyr732 robustly decreases its K and increases its V by disrupting its dimer formation. Mutation in c-Src phosphorylation site of either HK1 or HK2 remarkably abrogates the stimulating effects of c-Src on glycolysis, cell proliferation, migration, invasion, tumorigenesis and metastasis. Due to its lower K for glucose, HK1 rather than HK2 is required for tumour cell survival when glucose is scarce. Importantly, HK1-Y732 phosphorylation level remarkably correlates with the incidence and metastasis of various clinical cancers and may serve as a marker to predict metastasis risk of primary cancers.
Topics: Animals; Carcinogenesis; Cell Line, Tumor; Cell Proliferation; Enzyme Activation; Glucose; Glucose-6-Phosphate; Glycolysis; Heterografts; Hexokinase; Humans; Kinetics; Male; Metabolic Networks and Pathways; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Metastasis; Neoplasms; Phosphorylation; Protein Binding; Proto-Oncogene Proteins pp60(c-src); Tyrosine
PubMed: 28054552
DOI: 10.1038/ncomms13732 -
PloS One 2018Autophagy is systematically regulated by upstream factors and nutrients. Recent studies reported that telomerase and hexokinase 2 [HK2) regulate autophagy through mTOR...
Autophagy is systematically regulated by upstream factors and nutrients. Recent studies reported that telomerase and hexokinase 2 [HK2) regulate autophagy through mTOR and that telomerase has the capacity to bind to the HK2 promoter. However, the molecular linkage among telomerase, HK2, and autophagy is not fully understood. Here, we show that HK2 connects telomerase to autophagy. HK2 inhibition in HepG2 cells suppressed TERT-induced autophagy activation and further enhancement by glucose deprivation. The HK2 downstream factor mTOR was responsible for the TERT-induced autophagy activation under glucose deprivation, implying that TERT promotes autophagy through an HK2-mTOR pathway. TERC played a role similar to that of TERT, and simultaneous expression of TERT and TERC synergistically enhanced HK2 expression and autophagy. At the gene level, TERT bound to the HK2 promoter at a specific region harboring the telomerase-responsive sequence 'TTGGG.' Mutagenesis of TERC and the TERT-responsive element in the HK2 promoter revealed that TERC is required for the binding of TERT to the HK2 promoter. We demonstrate the existence of a telomerase-HK2-mTOR-autophagy axis and suggest that inhibition of the interaction between telomerase and the HK2 promoter diminishes glucose starvation-induced autophagy.
Topics: Autophagy; Glucose; Glycolysis; HeLa Cells; Hep G2 Cells; Hexokinase; Humans; Promoter Regions, Genetic; Telomerase
PubMed: 29462198
DOI: 10.1371/journal.pone.0193182 -
Journal of Neurochemistry Nov 2006Astrocytes play a well-established role in brain metabolism, being a key element in the capture of energetic compounds from the circulation and in their delivery to... (Review)
Review
Astrocytes play a well-established role in brain metabolism, being a key element in the capture of energetic compounds from the circulation and in their delivery to active neurons. Their metabolic status is affected in many pathological situations, such as gliomas, which are the most common brain tumors. This proliferative dysfunction is associated with changes in gap junctional communication, a property strongly developed in normal astrocytes studied both in vitro and in vivo. Here, we summarize and discuss the findings that have lead to the identification of a link between gap junctions, glucose uptake, and proliferation. Indeed, the inhibition of gap junctional communication is associated with an increase in glucose uptake due to a rapid change in the localization of both GLUT-1 and type I hexokinase. This effect persists due to the up-regulation of GLUT-1 and type I hexokinase and to the induction of GLUT-3 and type II hexokinase. In addition, cyclins D1 and D3 have been found to act as sensors of the inhibition of gap junctions and have been proposed to play the role of mediators in the mitogenic effect observed. Conversely, in C6 glioma cells, characterized by a low level of intercellular communication, an increase in gap junctional communication reduces glucose uptake by releasing type I and type II hexokinases from the mitochondria and decreases the exacerbated rate of proliferation due to the up-regulation of the Cdk inhibitors p21 and p27. Identification of the molecular actors involved in these pathways should allow the determination of potential therapeutic targets that could lead to the testing of alternative strategies to prevent, or at least slow down, the proliferation of glioma cells.
Topics: Animals; Astrocytes; Brain; Cell Communication; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p27; Energy Metabolism; Gap Junctions; Glucose; Glucose Transporter Type 1; Hexokinase; Humans; Neurons
PubMed: 16899068
DOI: 10.1111/j.1471-4159.2006.04088.x -
Eukaryotic Cell Jul 2010Fungi contain several hexokinases, which are involved either in sugar phosphorylation or in carbon source sensing. Glucose and fructose phosphorylations appear to rely...
Fungi contain several hexokinases, which are involved either in sugar phosphorylation or in carbon source sensing. Glucose and fructose phosphorylations appear to rely exclusively on glucokinase and hexokinase. Here, we characterized the catalytic glucokinase and hexokinase from the opportunistic human pathogen Aspergillus fumigatus and showed that both enzymes display different biochemical properties and play different roles during growth and development. Glucokinase efficiently activates glucose and mannose but activates fructose only to a minor extent. Hexokinase showed a high efficiency for fructose activation but also activated glucose and mannose. Transcript and activity determinations revealed high levels of glucokinase in resting conidia, whereas hexokinase was associated mainly with the mycelium. Consequentially, a glucokinase mutant showed delayed germination at low glucose concentrations, whereas colony growth was not overly affected. The deletion of hexokinase had only a minor impact on germination but reduced colony growth, especially on sugar-containing media. Transcript determinations from infected mouse lungs revealed the expression of both genes, indicating a contribution to virulence. Interestingly, a double-deletion mutant showed impaired growth not only on sugars but also on nonfermentable nutrients, and growth on gluconeogenic carbon sources was strongly suppressed in the presence of glucose. Furthermore, the glkA hxkA deletion affected cell wall integrity, implying that both enzymes contribute to the cell wall composition. Additionally, the absence of either enzyme deregulated carbon catabolite repression since mutants displayed an induction of isocitrate lyase activity during growth on glucose-ethanol medium. Therefore, both enzymes seem to be required for balancing carbon flux in A. fumigatus and are indispensable for growth under all nutritional conditions.
Topics: Animals; Aspergillosis; Aspergillus fumigatus; Biocatalysis; Carbon; Cell Wall; Gene Deletion; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Fungal; Glucokinase; Hexokinase; Humans; Lung; Mice; Mycelium; RNA, Messenger; Reverse Transcriptase Polymerase Chain Reaction; Spores, Fungal; Substrate Specificity; Sugar Phosphates; Trehalose
PubMed: 20453072
DOI: 10.1128/EC.00362-09 -
Bioscience Reports Feb 2018The high proliferation rate of tumor cells demands high energy and metabolites that are sustained by a high glycolytic flux known as the 'Warburg effect'. The activation...
The high proliferation rate of tumor cells demands high energy and metabolites that are sustained by a high glycolytic flux known as the 'Warburg effect'. The activation and further metabolism of glucose is initiated by hexokinase, a focal point of metabolic regulation. The human hexokinase 2 (HK2) is overexpressed in all aggressive tumors and predominantly found on the outer mitochondrial membrane, where interactions through its N-terminus initiates and maintains tumorigenesis. Here, we report the structure of HK2 in complex with glucose and glucose-6-phosphate (G6P). Structural and biochemical characterization of the mitochondrial conformation reveals higher conformational stability and slow protein unfolding rate () compared with the cytosolic conformation. Despite the active site similarity of all human hexokinases, the N-domain of HK2 is catalytically active but not in hexokinase 1 and 3. Helix-α that protrudes out of the N-domain to link it to the C-domain of HK2 is found to be important in maintaining the catalytic activity of the N-half. In addition, the N-domain of HK2 regulates the stability of the whole enzyme in contrast with the C-domain. Glucose binding enhanced the stability of the wild-type (WT) enzyme and the single mutant D657A of the C-domain, but it did not increase the stability of the D209A mutant of the N-domain. The interaction of HK2 with the mitochondria through its N-half is proposed to facilitate higher stability on the mitochondria. The identification of structural and biochemical differences between HK2 and other human hexokinase isozymes could potentially be used in the development of new anticancer therapies.
Topics: Catalysis; Catalytic Domain; Glucose; Glucose-6-Phosphate; Hexokinase; Humans; Isoenzymes; Kinetics; Mitochondria; Mitochondrial Membranes; Molecular Conformation; Mutation; Protein Conformation; Thermodynamics
PubMed: 29298880
DOI: 10.1042/BSR20171666 -
Cellular Immunology Mar 2023The adoptive transfer of ex vivo generated myeloid-derived suppressor cells (MDSCs) may be a promising therapeutic strategy for preventing allograft rejection after...
The adoptive transfer of ex vivo generated myeloid-derived suppressor cells (MDSCs) may be a promising therapeutic strategy for preventing allograft rejection after solid organ transplantation. Currently, the precise role of immune-metabolic pathways in the differentiation and function of MDSCs is not fully understood. Hexokinase 2 (HK2) is an isoform of hexokinase and is a key enzyme involved in the increased aerobic glycolysis of different immune cells during their activation and function. Here, we demonstrate that the addition of HK2 inhibitor 3-Bromopyruvic acid (3-BrPA) into traditional MDSCs induction system in vitro significantly promoted MDSCs production and enhanced their immunosuppressive function. Treatment with 3-BrPA increased the expression of MDSC-related immunosuppressive molecules, such as iNOS, Arg1, and CXCR2. Moreover, the adoptive transfer of 3-BrPA-treated MDSCs significantly prolonged the survival time of mouse heart allografts. This study provides a novel strategy to solve the problems of harvesting enough autologous cells for MDSC production from sick patients, and producing functionally enhanced MDSCs for preventing graft rejection and inducing tolerance.
Topics: Mice; Animals; Myeloid-Derived Suppressor Cells; Hexokinase; Immunosuppressive Agents; Organ Transplantation; Cell Differentiation
PubMed: 36774675
DOI: 10.1016/j.cellimm.2023.104688 -
European Journal of Biochemistry Jul 1990Increased contractile activity as induced by chronic low-frequency stimulation evoked in rat fast-twitch muscle an almost immediate increase in the ratio between...
Increased contractile activity as induced by chronic low-frequency stimulation evoked in rat fast-twitch muscle an almost immediate increase in the ratio between structure-bound and free hexokinase. In addition, an up to 14-fold rise in total hexokinase activity occurred after two weeks of stimulation indicating that glucose phosphorylation became a limiting step of glucose utilization under these conditions. The increase in hexokinase activity was transitory as prolonged stimulation led to a leveling off and steep decline with an apparent half-life of 2.5 days after three weeks of stimulation. The transient increase in glucose phosphorylating capacity can be explained by previous observations indicating that prolonged stimulation leads to a shift from a carbohydrate-based to a fatty-acid-based energy metabolism. Using an isozyme-specific sandwich ELISA, it was shown that both increases and decreases in total hexokinase activity were matched by corresponding changes in the amount of hexokinase isozyme II protein. Increases in both total hexokinase activity (3-4-fold) and hexokinase II protein content were also observed after denervation in rat fast-twitch muscle. In view of reports in the literature, it is suggested that the elevations in hexokinase II observed with increased contractile activity and denervation relate to enhanced glucose uptake and utilization.
Topics: Animals; Antibodies; Antibody Specificity; Denervation; Electric Stimulation; Enzyme-Linked Immunosorbent Assay; Evoked Potentials; Glucose; Hexokinase; Isoenzymes; Male; Models, Biological; Muscle Contraction; Muscle Proteins; Muscles; Phosphorylation; Rats; Rats, Inbred Strains
PubMed: 2379506
DOI: 10.1111/j.1432-1033.1990.tb19096.x -
Autophagy 2015Hexokinase is the first enzyme in the glycolytic pathway catalyzing the reaction in which glucose is phosphorylated into glucose-6-phosphate. Mammals possess 4 isoforms...
Hexokinase is the first enzyme in the glycolytic pathway catalyzing the reaction in which glucose is phosphorylated into glucose-6-phosphate. Mammals possess 4 isoforms of hexokinase; HK2 (hexokinase 2) is the predominant form in insulin-sensitive tissues such as adipocytes, as well as skeletal and cardiac muscle. In addition to its function in glucose metabolism, HK2 is associated with cardiomyocyte protection against mitochondrial-mediated apoptotic cell death; whether or not HK2 played a role in cardioprotective autophagy was yet to be discovered. However, in a recent study highlighted by a punctum in this issue of Autophagy, Roberts et al. addressed this possibility, uncovering a direct link between HK2, TORC1, and autophagy regulation.
Topics: Animals; Autophagy; Glycolysis; Hexokinase; Humans; Myocytes, Cardiac; Phosphorylation
PubMed: 26075875
DOI: 10.1080/15548627.2015.1042196 -
Nucleic Acids Research Jun 2022The aryl hydrocarbon receptor (AHR) regulates the expression of numerous genes in response to activation by agonists including xenobiotics. Although it is well...
The aryl hydrocarbon receptor (AHR) regulates the expression of numerous genes in response to activation by agonists including xenobiotics. Although it is well appreciated that environmental signals and cell intrinsic features may modulate this transcriptional response, how it is mechanistically achieved remains poorly understood. We show that hexokinase 2 (HK2) a metabolic enzyme fuelling cancer cell growth, is a transcriptional target of AHR as well as a modulator of its activity. Expression of HK2 is positively regulated by AHR upon exposure to agonists both in human cells and in mice lung tissues. Conversely, over-expression of HK2 regulates the abundance of many proteins involved in the regulation of AHR signalling and these changes are linked with altered AHR expression levels and transcriptional activity. HK2 expression also shows a negative correlation with AHR promoter methylation in tumours, and these tumours with high HK2 expression and low AHR methylation are associated with a worse overall survival in patients. In sum, our study provides novel insights into how AHR signalling is regulated which may help our understanding of the context-specific effects of this pathway and may have implications in cancer.
Topics: Animals; Hexokinase; Humans; Mice; Promoter Regions, Genetic; Receptors, Aryl Hydrocarbon; Signal Transduction; Xenobiotics
PubMed: 35609998
DOI: 10.1093/nar/gkac360 -
The Journal of Biological Chemistry 2021Hexokinase (HK) catalyzes the first step in glucose metabolism, making it an exciting target for the inhibition of tumor initiation and progression due to their elevated...
Hexokinase (HK) catalyzes the first step in glucose metabolism, making it an exciting target for the inhibition of tumor initiation and progression due to their elevated glucose metabolism. The upregulation of hexokinase-2 (HK2) in many cancers and its limited expression in normal tissues make it a particularly attractive target for the selective inhibition of cancer growth and the eradication of tumors with limited side effects. The design of such safe and effective anticancer therapeutics requires the development of HK2-specific inhibitors that will not interfere with other HK isozymes. As HK2 is unique among HKs in having a catalytically active N-terminal domain (NTD), we have focused our attention on this region. We previously found that NTD activity is affected by the size of the linker helix-α that connects the N- and C-terminal domains of HK2. Three nonactive site residues (D447, S449, and K451) at the beginning of the linker helix-α have been found to regulate the NTD activity of HK2. Mutation of these residues led to increased dynamics, as shown via hydrogen deuterium exchange analysis and molecular dynamic simulations. D447A contributed the most to the enhanced dynamics of the NTD, with reduced calorimetric enthalpy of HK2. Similar residues exist in the C-terminal domain (CTD) but are unnecessary for HK1 and HK2 activity. Thus, we postulate these residues serve as a regulatory site for HK2 and may provide new directions for the design of anticancer therapeutics that reduce the rate of glycolysis in cancer through specific inhibition of HK2.
Topics: Antineoplastic Agents; Catalysis; Cell Line, Tumor; Enzyme Inhibitors; Enzyme Stability; Glycolysis; Hexokinase; Humans; Kinetics; Protein Domains; Substrate Specificity
PubMed: 33187984
DOI: 10.1074/jbc.RA120.015293