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Glucose starvation activates AMPK via an AMP/ADP-independent mechanism that involves fructose-1,6-bisphosphate and aldolase. New insights into AMPK activation AMPK is a central regulator of metabolic homeostasis, and its dysfunction may result in various diseases including diabetes, obesity, and cancer. AMPK is known to be activated under stressful conditions, including glucose starvation. It has been assumed that upon glucose deprivation AMPK activation occurs in the canonical AMP/ADP-dependent manner, with reduced metabolism of glucose causing falling ATP and increasing AMP and ADP. Here, Sheng-Cai Lin and colleagues show that this is not the case, and that glucose starvation activates AMPK via a different route, in an AMP/ADP-independent manner. During glycolysis, glucose is converted to fructose-1,6-bisphosphate (FBP), which is then processed by FBP aldolases. The authors show that the absence of glucose results in a reduction of FBP-bound aldolase, which triggers LKB1 phosphorylation and activation of AMPK. This study thus uncovers FBP as the critical metabolite that signals glucose availability and FBP aldolases as the sensors that relay the information to AMPK. The major energy source for most cells is glucose, from which ATP is generated via glycolysis and/or oxidative metabolism. Glucose deprivation activates AMP-activated protein kinase (AMPK) 1 , but it is unclear whether this activation occurs solely via changes in AMP or ADP, the classical activators of AMPK 2 , 3 , 4 , 5 . Here, we describe an AMP/ADP-independent mechanism that triggers AMPK activation by sensing the absence of fructose-1,6-bisphosphate (FBP), with AMPK being progressively activated as extracellular glucose and intracellular FBP decrease. When unoccupied by FBP, aldolases promote the formation of a lysosomal complex containing at least v-ATPase, ragulator, axin, liver kinase B1 (LKB1) and AMPK, which has previously been shown to be required for AMPK activation 6 , 7 . Knockdown of aldolases activates AMPK even in cells with abundant glucose, whereas the catalysis-defective D34S aldolase mutant, which still binds FBP, blocks AMPK activation. Cell-free reconstitution assays show that addition of FBP disrupts the association of axin and LKB1 with v-ATPase and ragulator. Importantly, in some cell types AMP/ATP and ADP/ATP ratios remain unchanged during acute glucose starvation, and intact AMP-binding sites on AMPK are not required for AMPK activation. These results establish that aldolase, as well as being a glycolytic enzyme, is a sensor of glucose availability that regulates AMPK.

Yeast and cancer cells share the unusual characteristic of favoring fermentation of sugar over respiration. We now reveal an evolutionary conserved mechanism linking fermentation to activation of Ras, a major regulator of cell proliferation in yeast and mammalian cells, and prime proto-oncogene product. A yeast mutant ( tps1∆ ) with overactive influx of glucose into glycolysis and hyperaccumulation of Fru1,6bisP, shows hyperactivation of Ras, which causes its glucose growth defect by triggering apoptosis. Fru1,6bisP is a potent activator of Ras in permeabilized yeast cells, likely acting through Cdc25. As in yeast, glucose triggers activation of Ras and its downstream targets MEK and ERK in mammalian cells. Biolayer interferometry measurements show that physiological concentrations of Fru1,6bisP stimulate dissociation of the pure Sos1/H-Ras complex. Thermal shift assay confirms direct binding to Sos1, the mammalian ortholog of Cdc25. Our results suggest that the Warburg effect creates a vicious cycle through Fru1,6bisP activation of Ras, by which enhanced fermentation stimulates oncogenic potency. Yeast and cancer cells both favor sugar fermentation in aerobic conditions. Here the authors describe a conserved mechanism from yeast to mammals where the glycolysis intermediate fructose-1,6-bisphosphate binds Cdc25/Sos1 and couples increased glycolytic flux to increased Ras proto-oncoprotein activity.

Metabolic programming and mitochondrial dynamics along with T cell differentiation affect T cell fate and memory development; however, how to control metabolic reprogramming and mitochondrial dynamics in T cell memory development is unclear. Here, we provide evidence that the SUMO protease SENP1 promotes T cell memory development via Sirt3 deSUMOylation. SENP1-Sirt3 signalling augments the deacetylase activity of Sirt3, promoting both OXPHOS and mitochondrial fusion. Mechanistically, SENP1 activates Sirt3 deacetylase activity in T cell mitochondria, leading to reduction of the acetylation of mitochondrial metalloprotease YME1L1. Consequently, deacetylation of YME1L1 suppresses its activity on OPA1 cleavage to facilitate mitochondrial fusion, which results in T cell survival and promotes T cell memory development. We also show that the glycolytic intermediate fructose-1,6-bisphosphate (FBP) as a negative regulator suppresses AMPK-mediated activation of the SENP1-Sirt3 axis and reduces memory development. Moreover, glucose limitation reduces FBP production and activates AMPK during T cell memory development. These data show that glucose limitation activates AMPK and the subsequent SENP1-Sirt3 signalling for T cell memory development. Memory T cells are particularly reliant on fatty acid oxidation as a source of energy. Here the authors show this reliance is controlled by AMPK sensing of glucose deprivation that triggers SENP1-Sirt3 signalling, driving fatty acid oxidation and memory differentiation in T cells via deacetylation of YME1L1 to induce mitochondrial fusion.

TIGAR (TP53-induced glycolysis and apoptosis regulator) is the downstream target gene of p53, contains a functional sequence similar to 6-phosphofructose kinase/fructose-2, 6-bisphosphatase (PFKFB) bisphosphatase domain. TIGAR is mainly located in the cytoplasm; in response to stress, TIGAR is translocated to nucleus and organelles, including mitochondria and endoplasmic reticulum to regulate cell function. P53 family members (p53, p63, and p73), some transcription factors (SP1 and CREB), and noncoding miRNAs (miR-144, miR-885-5p, and miR-101) regulate the transcription of TIGAR. TIGAR mainly functions as fructose-2,6-bisphosphatase to hydrolyze fructose-1,6-diphosphate and fructose-2,6-diphosphate to inhibit glycolysis. TIGAR in turn facilitates pentose phosphate pathway flux to produce nicotinamide adenine dinucleotide phosphate (NADPH) and ribose, thereby promoting DNA repair, and reducing intracellular reactive oxygen species. TIGAR thus maintains energy metabolism balance, regulates autophagy and stem cell differentiation, and promotes cell survival. Meanwhile, TIGAR also has a nonenzymatic function and can interact with retinoblastoma protein, protein kinase B, nuclear factor-kappa B, hexokinase 2, and ATP5A1 to mediate cell cycle arrest, inflammatory response, and mitochondrial protection. TIGAR might be a potential target for the prevention and treatment of cardiovascular and neurological diseases, as well as cancers.

The enzyme fructose-bisphosphate aldolase occupies a central position in glycolysis and gluconeogenesis pathways. Beyond its housekeeping role in metabolism, fructose-bisphosphate aldolase has been involved in additional functions and is considered as a potential target for drug development against pathogenic bacteria. Here, we address the role of fructose-bisphosphate aldolase in the bacterial pathogen Francisella novicida. We demonstrate that fructose-bisphosphate aldolase is important for bacterial multiplication in macrophages in the presence of gluconeogenic substrates. In addition, we unravel a direct role of this metabolic enzyme in transcription regulation of genes katG and rpoA, encoding catalase and an RNA polymerase subunit, respectively. We propose a model in which fructose-bisphosphate aldolase participates in the control of host redox homeostasis and the inflammatory immune response.

Coxiella burnetii is a bacterial pathogen that replicates within host cells by establishing a membrane-bound niche called the Coxiella-containing vacuole. Biogenesis of this compartment requires effectors of its Dot/Icm type IV secretion system. A large cohort of such effectors has been identified, but the function of most of them remain elusive. Here, by a cell-based functional screening, we identified the effector Cbu0513 (designated as CinF) as an inhibitor of NF-κB signaling. CinF is highly similar to a fructose-1,6-bisphosphate (FBP) aldolase/phosphatase present in diverse bacteria. Further study reveals that unlike its ortholog from Sulfolobus tokodaii, CinF does not exhibit FBP phosphatase activity. Instead, it functions as a protein phosphatase that specifically dephosphorylates and stabilizes IκBα. The IκBα phosphatase activity is essential for the role of CinF in C. burnetii virulence. Our results establish that C. burnetii utilizes a protein adapted from sugar metabolism to subvert host immunity.

While fructose-1,6-diphosphate (FDP) has been clinically applied in ischemic conditions, its comprehensive effects on hemostasis remain to be fully elucidated. This in vitro study examined the influence of FDP on coagulation and platelet function using thromboelastography (TEG) across three platforms, coagulation factor assays, and platelet aggregation tests over a concentration range of 0–6 mg/mL. The results revealed that FDP concentration-dependently prolonged TEG R-time ( P  < 0.01), with an increase of 21.8–48.3% at the clinically relevant concentration of 3.71 mg/mL. Strong inverse correlations were observed between FDP concentration and the activities of factors V, VII, IX, XI, and XII ( r  = − 0.989 to − 0.997, P  < 0.001), whereas factors II, VIII, and X remained unaltered. Furthermore, FDP significantly inhibited platelet aggregation ( P  < 0.001), nearly abolishing epinephrine- and ADP-induced aggregation at 6 mg/mL under unbuffered conditions. pH-control experiments confirmed that the anticoagulant effects were FDP-specific, while the antiplatelet effects were primarily mediated by FDP with a partial pH-dependent component. These findings demonstrate that FDP possesses dual anticoagulant and antiplatelet properties through selective inhibition of coagulation initiation factors and broad suppression of platelet responsiveness, suggesting potential implications for clinical use in high-risk populations and warranting further investigation.

Necrotic enteritis is an economically important poultry disease caused by the bacterium Clostridium perfringens. There are currently no necrotic enteritis vaccines commercially available for use in broiler birds, the most important target population. Salmonella-vectored vaccines represent a convenient and effective option for controlling this disease. We used a single attenuated Salmonella vaccine strain, engineered to lyse within the host, to deliver up to three C. perfringens antigens. Two of the antigens were toxoids, based on C. perfringens α-toxin and NetB toxin. The third antigen was fructose-1,6-bisphosphate aldolase (Fba), a metabolic enzyme with an unknown role in virulence. Oral immunization with a single Salmonella vaccine strain producing either Fba, α-toxoid and NetB toxoid, or all three antigens, was immunogenic, inducing serum, cellular and mucosal responses against Salmonella and the vectored C. perfringens antigens. All three vaccine strains were partially protective against virulent C. perfringens challenge. The strains delivering Fba only or all three antigens provided the best protection. We also demonstrate that both toxins and Fba are present on the C. perfringens cell surface. The presence of Fba on the cell surface suggests that Fba may function as an adhesin.

Fructose-bisphosphate aldolase A (ALDOA) is a key enzyme in glycolysis and is responsible for catalyzing the reversible conversion of fructose-1,6-bisphosphate to glyceraldehydes-3-phosphate and dihydroxyacetone phosphate. ALDOA contributes to various cellular functions such as muscle maintenance, regulation of cell shape and mobility, striated muscle contraction, actin filament organization and ATP biosynthetic process. Here, we reported that ALDOA is a highly expressed in lung squamous cell carcinoma (LSCC) and its expression level is correlated with LSCC metastasis, grades, differentiation status and poor prognosis. Depletion of ALDOA expression in the lung squamous carcinoma NCI-H520 cells reduces the capabilities of cell motility and tumorigenesis. These data suggest that ALDOA could be a potential marker for LSCC metastasis and a therapeutic target for drug development.

Muscle fructose-1,6-bisphosphate aldolase (ALDOA) is among the most abundant glycolytic enzymes in all cancer cells. Here, we show that the enzyme plays a previously unknown and critical role in a cancer cell survival. Simultaneous inhibition of ALDOA activity and interaction with F-actin cytoskeleton using ALDOA slow-binding inhibitor UM0112176 leads to a rapid cofilin-dependent loss of F-actin stress fibers which is associated with elevated ROS production, inhibition of ATP synthesis, increase in calcium levels, caspase activation and arrested cellular proliferation. These effects can be reproduced by silencing of ALDOA. The mechanism of pharmacological action is, however, independent of the catalytic function of the enzyme, specific to cancer cells, and is most deleterious to cells undergoing the epithelial–mesenchymal transition, a process facilitating cancer cell invasion. Our results demonstrate that the overabundance of ALDOA in cancer cells is associated with its moonlighting rather than catalytic functions. This may have significant implications for development of novel broad-based anti-cancer therapies.