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Objective: Stable isotope studies have shown that hepatic de novo lipogenesis (DNL) plays an important role in the pathogenesis of intrahepatic lipid (IHL) deposition. Furthermore, previous research has demonstrated that fructose 1-phosphate (F1P) not only serves as a substrate for DNL, but also acts as a signalling metabolite that stimulates DNL from glucose. The aim of this study was to elucidate the mediators of F1P-stimulated DNL, with special focus on two key regulators of intrahepatic glucose metabolism, i.e., glucokinase regulatory protein (GKRP) and carbohydrate response element binding protein (ChREBP)., Methods: Aldolase B deficient mice (Aldob -/- ), characterized by hepatocellular F1P accumulation, enhanced DNL, and hepatic steatosis, were either crossed with GKRP deficient mice (Gckr -/- ) or treated with short hairpin RNAs directed against hepatic ChREBP., Results: Aldob -/- mice showed higher rates of de novo palmitate synthesis from glucose when compared to wildtype mice (p < 0.001). Gckr knockout reduced de novo palmitate synthesis in Aldob -/- mice (p = 0.017), without affecting the hepatic mRNA expression of enzymes involved in DNL. In contrast, hepatic ChREBP knockdown normalized the hepatic mRNA expression levels of enzymes involved in DNL and reduced fractional DNL in Aldob -/- mice (p < 0.05). Of interest, despite downregulation of DNL in response to Gckr and ChREBP attenuation, no reduction in intrahepatic triglyceride levels was observed., Conclusions: Both GKRP and ChREBP mediate F1P-stimulated DNL in aldolase B deficient mice. Further studies are needed to unravel the role of GKRP and hepatic ChREBP in regulating IHL accumulation in aldolase B deficiency., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Despite being the leading cause of lung cancer-related deaths, the underlying molecular mechanisms driving metastasis progression are still not fully understood. Transfer RNA-derived fragments (tRFs) have been implicated in various biological processes in cancer. However, the role of tRFs in lung adenocarcinoma (LUAD) remains unclear. Our study identified a tRF, tRF-Val-CAC-024, associated with the high-risk component of LUAD, through validation using 3 cohorts. Our findings demonstrated that tRF-Val-CAC-024 acts as an oncogene in LUAD. Mechanistically, tRF-Val-CAC-024 was revealed to bind to aldolase A (ALDOA) dependent on Q125/E224 and promote the oligomerization of ALDOA, resulting in increased enzyme activity and enhanced aerobic glycolysis in LUAD cells. Additionally, we provide preliminary evidence of its potential clinical value by investigating the therapeutic effects of tRF-Val-CAC-024 antagomir-loaded lipid nanoparticles (LNPs) in cell-line-derived xenograft models. These results could enhance our understanding of the regulatory mechanisms of tRFs in LUAD and provide a potential therapeutic target., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Peptostreptococcus stomatis (P. stomatis) is enriched in colorectal cancer (CRC), but its causality and translational implications in CRC are unknown. Here, we show that P. stomatis accelerates colonic tumorigenesis in Apc Min/+ and azoxymethane/dextran sodium sulfate (AOM-DSS) models by inducing cell proliferation, suppressing apoptosis, and impairing gut barrier function. P. stomatis adheres to CRC cells through its surface protein fructose-1,6-bisphosphate aldolase (FBA) that binds to the integrin α6/β4 receptor on CRC cells, leading to the activation of ERBB2 and the downstream MEK-ERK-p90 cascade. Blockade of the FBA-integrin α6/β4 abolishes ERBB2-mitogen-activated protein kinase (MAPK) activation and the protumorigenic effect of P. stomatis. P. stomatis-driven ERBB2 activation bypasses receptor tyrosine kinase (RTK) blockade by EGFR inhibitors (cetuximab, erlotinib), leading to drug resistance in xenograft and spontaneous CRC models of KRAS-wild-type CRC. P. stomatis also abrogates BRAF inhibitor (vemurafenib) efficacy in BRAF V600E -mutant CRC xenografts. Thus, we identify P. stomatis as an oncogenic bacterium and a contributory factor for non-responsiveness to RTK inhibitors in CRC., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Fructose bisphosphate aldolases (FBAs) catalyze the reversible cleavage of fructose 1,6-bisphosphate into dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. We analyzed two previously uncharacterized cytosolic Arabidopsis FBAs, AtFBA4 and AtFBA5. Based on a recent report, we examined the interaction of AtFBA4 with calmodulin (CaM)-like protein 11 (AtCML11). AtFBA4 did not bind AtCML11; however, we found that CaM bound AtFBA5 in a Ca 2+ -dependent manner with high specificity and affinity (K D  ~ 190 nm) and enhanced its stability. AtFBA4 and AtFBA5 exhibited Michaelis-Menten kinetics with K m and V max values of 180 μm and 4.9 U·mg -1 for AtFBA4, and 6.0 μm and 0.30 U·mg -1 for AtFBA5, respectively. The flavonoid morin inhibited both isozymes. Our study suggests that Ca 2+ signaling and flavanols may influence plant glycolysis/gluconeogenesis., (© 2024 The Author(s). FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Hereditary fructose intolerance (HFI) is an autosomal recessive metabolic disease associated with a mutation in the aldolase B gene on chromosome 9q31. In this study, we generated a human-induced pluripotent stem cell (hiPSC) line, FDCHi015-A, from peripheral blood mononuclear cells (PBMCs) of a patient carrying the compound heterozygous mutations c.360&#95;364delCAAA and c.1013C > T in exons 4 and 9 of the ALDOB gene, respectively. The iPSCs with the confirmed patient-specific mutation demonstrate pluripotency markers expression, a normal karyotype, and the ability to differentiate into derivatives of three germ layers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Fructose 1,6-bisphosphate aldolase (FBA) is a pivotal enzyme, which plays a critical role in fixing CO 2 through the process of in the Calvin cycle. In this study, a comprehensive exploration of the FBA family genes in moso bamboo (Phyllostachys edulis) was conducted by the bioinformatics and biological analyses. A total of nine FBA genes (PeFBA1-PeFBA9) were identified in the moso bamboo genome. The expression patterns of PeFBAs across diverse tissues of moso bamboo suggested that they have multifaceted functionality. Notably, PeFBA8 might play an important role in regulating photosynthetic carbon metabolism. Co-expression and cis-element analyses demonstrated that PeFBA8 was regulated by a photosynthetic regulatory transcription factor (PeGLK1), which was confirmed by yeast one-hybrid and dual-luciferase assays. In-planta gene editing analysis revealed that the edited PeFBA8 mutants displayed compromised photosynthetic functionality, characterized by reduced electron transport rate and impaired photosystem I, leading to decreased photosynthesis rate overall, compared to the unedited control. The recombinant protein of PeFBA8 from prokaryotic expression exhibited enzymatic catalytic function. The findings suggest that the expression of PeFBA8 can affect photosynthetic efficiency of moso bamboo leaves, which underlines the potential of leveraging PeFBA8's regulatory mechanism to breed bamboo varieties with enhanced carbon fixation capability., Competing Interests: Declaration of competing interest No conflict of interest exits in the submission of this manuscript, and it is approved by all authors for publication. I would like to declare on behalf of my co-authors that the work described is original research that has not been published previously, and not under consideration for publication elsewhere, in whole or in part. All the authors listed have approved the manuscript that is enclosed., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The fructose-1,6-bisphosphate aldolase ( FBA ) gene family exists in higher plants, with the genes of this family playing significant roles in plant growth and development, as well as response to abiotic stresses. However, systematic reports on the FBA gene family and its functions in cucumber are lacking. In this study, we identified five cucumber FBA genes, named CsFBA1 - 5 , that are distributed randomly across chromosomes. Phylogenetic analyses involving these cucumber FBAs, alongside eight Arabidopsis FBA proteins and eight tomato FBA proteins, were conducted to assess their homology. The CsFBAs were grouped into two clades. We also analyzed the physicochemical properties, motif composition, and gene structure of the cucumber FBAs. This analysis highlighted differences in the physicochemical properties and revealed highly conserved domains within the CsFBA family. Additionally, to explore the evolutionary relationships of the CsFBA family further, we constructed comparative syntenic maps with Arabidopsis and tomato, which showed high homology but only one segmental duplication event within the cucumber genome. Expression profiles indicated that the CsFBA gene family is responsive to various abiotic stresses, including low temperature, heat, and salt. Taken together, the results of this study provide a theoretical foundation for understanding the evolution of and future research into the functional characterization of cucumber FBA genes during plant growth and development.

Hereditary fructose intolerance (HFI) is a painful and potentially lethal genetic disease caused by a mutation in aldolase B resulting in accumulation of fructose-1-phosphate (F1P). No cure exists for HFI and treatment is limited to avoid exposure to fructose and sugar. Using aldolase B deficient mice, here we identify a yet unrecognized metabolic event activated in HFI and associated with the progression of the disease. Besides the accumulation of F1P, here we show that the activation of the purine degradation pathway is a common feature in aldolase B deficient mice exposed to fructose. The purine degradation pathway is a metabolic route initiated by adenosine monophosphate deaminase 2 (AMPD2) that regulates overall energy balance. We demonstrate that very low amounts of fructose are sufficient to activate AMPD2 in these mice via a phosphate trap. While blocking AMPD2 do not impact F1P accumulation and the risk of hypoglycemia, its deletion in hepatocytes markedly improves the metabolic dysregulation induced by fructose and corrects fat and glycogen storage while significantly increasing the voluntary tolerance of these mice to fructose. In summary, we provide evidence for a critical pathway activated in HFI that could be targeted to improve the metabolic consequences associated with fructose consumption., (© 2024. The Author(s).)

Prior studies have emphasized a bioenergetic crisis in the retinal pigment epithelium (RPE) as a critical factor in the development of age-related macular degeneration (AMD). The isoforms Fructose-1,6-bisphosphate aldolase C (ALDOC) and pyruvate kinase M2 (PKM2) have been proposed to play a role in AMD pathogenesis. While PKM2 and ALDOC are crucial for aerobic glycolysis in the neural retina, they are not as essential for the RPE. In this study, we examined the expression and activity of PKM2 and ALDOC in both young and aged RPE cells, as well as in the retina and RPE tissue of mice, including an experimentally induced AMD mouse model. Our findings reveal an upregulation in PKM2 and ALDOC expression, accompanied by increased pyruvate kinase activity, in the aged and AMD mouse RPE. Conversely, there is a decrease in ALDOC expression but an increase in PKM2 expression and pyruvate kinase activity in the aged and AMD retina. Overall, our study indicates that aged and AMD RPE cells tend to favor aerobic glycolysis, while this tendency is diminished in the aged and AMD retina. These results underscore the significance of targeting PKM2 and ALDOC in the RPE as a promising therapeutic approach to address the bioenergetic crisis and prevent vision loss in AMD.

Rationale : CD8 + T cells undergo a series of metabolic reprogramming processes during their activation and proliferation, including increased glycolysis, decreased aerobic oxidation of sugars, increased amino acid metabolism and increased protein synthesis. However, it is still unclear what factors regulate these metabolic reprogramming processes in CD8 + T cells in the tumor immune microenvironment. Methods : T cell chromobox protein 4 (CBX4) knock-out mice models were used to determine the role of CBX4 in CD8 + T cells on the tumor immune microenvironment and tumor progression. Flow cytometry, Cut-Tag qPCR, Chip-seq, immunoprecipitation, metabolite detection, lentivirus infection and adoptive T cells transfer were performed to explore the underlying mechanisms of CBX4 knock-out in promoting CD8+ T cell activation and inhibiting tumor growth. Results : We found that CBX4 expression was induced in tumor-infiltrating CD8 + T cells and inhibited CD8 + T cell function by regulating glucose metabolism in tumor tissue. Mechanistically, CBX4 increases the expression of the metabolism-associated molecule aldolase B (Aldob) through sumoylation of trans-acting transcription factor 1 (SP1) and Krüppel-like factor 3 (KLF3). In addition, Aldob inhibits glycolysis and ATP synthesis in T cells by reducing the phosphorylation of the serine/threonine protein kinase (Akt) and ultimately suppresses CD8 + T cell function. Significantly, knocking out CBX4 may improve the efficacy of anti-PD-1 therapy by enhancing the function of CD8 + T cells in the tumor microenvironment. Conclusion : CBX4 is involved in CD8 + T cell metabolic reprogramming and functional persistence in tumor tissues, and serves as an inhibitor in CD8 + T cells' glycolysis and effector function., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

The increasing availability of experimental and computational protein structures entices their use for function prediction. Here we develop an automated procedure to identify enzymes involved in metabolic reactions by assessing substrate conformations docked to a library of protein structures. By screening AlphaFold-modeled vitamin B6-dependent enzymes, we find that a metric based on catalytically favorable conformations at the enzyme active site performs best (AUROC Score=0.84) in identifying genes associated with known reactions. Applying this procedure, we identify the mammalian gene encoding hydroxytrimethyllysine aldolase (HTMLA), the second enzyme of carnitine biosynthesis. Upon experimental validation, we find that the top-ranked candidates, serine hydroxymethyl transferase (SHMT) 1 and 2, catalyze the HTMLA reaction. However, a mouse protein absent in humans (threonine aldolase; Tha1) catalyzes the reaction more efficiently. Tha1 did not rank highest based on the AlphaFold model, but its rank improved to second place using the experimental crystal structure we determined at 2.26 Å resolution. Our findings suggest that humans have lost a gene involved in carnitine biosynthesis, with HTMLA activity of SHMT partially compensating for its function., (© 2024. The Author(s).)

Zinc is required for many critical processes, including intermediary metabolism. In Saccharomyces cerevisiae, the Zap1 activator regulates the transcription of ∼80 genes in response to Zn supply. Some Zap1-regulated genes are Zn transporters that maintain Zn homeostasis, while others mediate adaptive responses that enhance fitness. One adaptive response gene encodes the 2-cysteine peroxiredoxin Tsa1, which is critical to Zn-deficient (ZnD) growth. Depending on its redox state, Tsa1 can function as a peroxidase, a protein chaperone, or a regulatory redox sensor. In a screen for possible Tsa1 regulatory targets, we identified a mutation (cdc19 S492A ) that partially suppressed the tsa1Δ growth defect. The cdc19 S492A mutation reduced activity of its protein product, pyruvate kinase isozyme 1 (Pyk1), implicating Tsa1 in adapting glycolysis to ZnD conditions. Glycolysis requires activity of the Zn-dependent enzyme fructose-bisphosphate aldolase 1, which was substantially decreased in ZnD cells. We hypothesized that in ZnD tsa1Δ cells, the loss of a compensatory Tsa1 regulatory function causes depletion of glycolytic intermediates and restricts dependent amino acid synthesis pathways, and that the decreased activity of Pyk1 S492A counteracted this depletion by slowing the irreversible conversion of phosphoenolpyruvate to pyruvate. In support of this model, supplementing ZnD tsa1Δ cells with aromatic amino acids improved their growth. Phosphoenolpyruvate supplementation, in contrast, had a much greater effect on growth rate of WT and tsa1Δ ZnD cells, indicating that inefficient glycolysis is a major factor limiting yeast growth. Surprisingly however, this restriction was not primarily due to low fructose-bisphosphate aldolase 1 activity, but instead occurs earlier in glycolysis., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Published by Elsevier Inc.)

Hepatocellular carcinoma (HCC) is one of the malignancies with the worst prognosis worldwide, in the occurrence and development of which glycolysis plays a central role. This study uncovered a mechanism by which ZNF692 regulates ALDOA-dependent glycolysis in HCC cells. RT-qPCR and western blotting were used to detect the expression of ZNF692, KAT5, and ALDOA in HCC cell lines and a normal liver cell line. The influences of transfection-induced alterations in the expression of ZNF692, KAT5, and ALDOA on the functions of HepG2 cells were detected by performing MTT, flow cytometry, Transwell, cell scratch, and colony formation assays, and the levels of glucose and lactate were determined using assay kits. ChIP and luciferase reporter assays were conducted to validate the binding of ZNF692 to the KAT5 promoter, and co-IP assays to detect the interaction between KAT5 and ALDOA and the acetylation of ALDOA. ZNF692, KAT5, and ALDOA were highly expressed in human HCC samples and cell lines, and their expression levels were positively correlated in HCC. ZNF692, ALDOA, or KAT5 knockdown inhibited glycolysis, proliferation, invasion, and migration and promoted apoptosis in HepG2 cells. ZNF692 bound to the KAT5 promoter and promoted its activity. ALDOA acetylation levels were elevated in HCC cell lines. KAT5 bound to ALDOA and catalyzed ALDOA acetylation. ALDOA or KAT5 overexpression in the same time of ZNF692 knockdown, compared to ZNF692 knockdown only, stimulated glycolysis, proliferation, invasion, and migration and reduced apoptosis in HepG2 cells. ZNF692 promotes the acetylation modification and protein expression of ALDOA by catalyzing KAT5 transcription, thereby accelerating glycolysis to drive HCC cell development., (© 2024. The Author(s).)

The availability of genome-wide transcriptomic and proteomic datasets is ever-increasing and often not used beyond initial publication. Here, we applied module-based coexpression network analysis to a comprehensive catalog of 35 mouse genome-wide liver expression datasets (encompassing more than 3800 mice) with the goal of identifying and validating unknown genes involved in cholesterol metabolism. From these 35 datasets, we identified a conserved module of genes enriched with cholesterol biosynthetic genes. Using a systematic approach across the 35 datasets, we identified three genes (Rdh11, Echdc1, and Aldoc) with no known role in cholesterol metabolism. We then performed functional validation studies and show that each gene is capable of regulating cholesterol metabolism. For the glycolytic gene, Aldoc, we demonstrate that it contributes to de novo cholesterol biosynthesis and regulates cholesterol and triglyceride levels in mice. As Aldoc is located within a genome-wide significant genome-wide association studies locus for human plasma cholesterol levels, our studies establish Aldoc as a causal gene within this locus. Through our work, we develop a framework for leveraging mouse genome-wide liver datasets for identifying and validating genes involved in cholesterol metabolism., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Lung cancer has a high incidence rate and requires more effective treatment strategies and drug options for clinical patients. EGFR is a common genetic alteration event in lung cancer that affects patient survival and drug strategy. Our study discovered aberrant aldolase A (ALDOA) expression and dysfunction in lung cancer patients with EGFR mutations. In addition to investigating relevant metabolic processes like glucose uptake, lactate production, and ATPase activity, we examined multi-omics profiles (transcriptomics, proteomics, and pull-down assays). It was observed that phosphodiesterase 3A (PDE3A) enzyme and ALDOA exhibit correlation, and furthermore, they impact M2 macrophage polarization through β-catenin and downstream ID3. In addition to demonstrating the aforementioned mechanism of action, our experiments discovered that the PDE3 inhibitor trequinsin has a substantial impact on lung cancer cell lines with EGFR mutants. The trequinsin medication was found to decrease the M2 macrophage polarization status and several cancer phenotypes, in addition to transduction. These findings have potential prognostic and therapeutic applications for clinical patients with EGFR mutation and lung cancer., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objectives: To research the associations between fructose-bisphosphate aldolase B (ALDOB) gene polymorphisms and intrahepatic cholestasis of pregnancy (ICP) risk., Material and Methods: Whole-genome sequencing (WGS) was performed to detect ALDOB polymorphisms. Five web-available tools were employed to predict the effect of the site variant on the protein. Protein structure comparisons between the reference and ALDOB-modified samples were performed by SWISS-MODEL and Chimera 1.14rc, respectively., Results: We identified 28 genetic variants in the ALDOB gene. When the cut-off value of minor allele frequency (MAF) of loci was 0.001 in four databases, five missense variants, including rs747604233, rs759204107, rs758242037, rs371526091 and rs77718928, were reserved for subsequent analysis. These variants were absent from the 1029 control individuals. The influence of all five variants on protein function was predicted to be damaging by the abovementioned five prediction software programs. Bioinformatics analysis demonstrated that these five missense variants were highly conserved among vertebrates. Compared to the wild-type protein structure, all five mutated protein structures showed a slight change in the chemical bond lengths of the enzyme activity domains. The combined clinical data indicate that the variant group had a significantly older age (p = 0.038), a higher level of indirect bilirubin (IDBIL, p = 0.033), and lower counts of white blood cells (WBCs, p = 7.38E-05) and platelets (PLTs, p = 0.018) than the wild-type group., Conclusions: This is the first study to examine the associations between ALDOB polymorphisms and ICP disease in 249 Chinese patients with ICP. Our present study expands the understanding of the pathogenesis of ICP.

Radioresistance is the major obstacle that affects the efficacy of radiotherapy which is an important treatment for cervical cancer. By analyzing the databases, we found that aldolase A (ALDOA), which is a key enzyme in metabolic reprogramming, has a higher expression in cervical cancer patients and is associated with poor prognosis. We detected the expression of ALDOA in the constructed cervical cancer radioresistance (RR) cells by repetitive irradiation and found that it was upregulated compared to the control cells. Functional assays were conducted and the results showed that the knockdown of ALDOA in cervical cancer RR cells inhibited the proliferation, migration, and clonogenic abilities by regulating the cell glycolysis. In addition, downregulation of ALDOA enhanced radiation-induced apoptosis and DNA damage by causing G2/M phase arrest and further promoted radiosensitivity of cervical cancer cells. The functions of ALDOA in regulating tumor radiosensitivity were also verified by the mouse tumor transplantation model in vivo . Therefore, our study provides new insights into the functions of ALDOA in regulating the efficacy of radiotherapy and indicates that ALDOA might be a promising target for enhancing radiosensitivity in treating cervical cancer patients.

Background: LIPH, a membrane-associated phosphatidic acid-selective phospholipase A1a, can produce LPA (Lysophosphatidic acid) from PA (Phosphatidic acid) on the outer leaflet of the plasma membrane. It is well known that LIPH dysfunction contributes to lipid metabolism disorder. Previous study shows that LIPH was found to be a potential gene related to poor prognosis with pancreatic ductal adenocarcinoma (PDAC). However, the biological functions of LIPH in PDAC remain unclear., Methods: Cell viability assays were used to evaluate whether LIPH affected cell proliferation. RNA sequencing and immunoprecipitation showed that LIPH participates in tumor glycolysis by stimulating LPA/LPAR axis and maintaining aldolase A (ALDOA) stability in the cytosol. Subcutaneous, orthotopic xenograft models and patient-derived xenograft PDAC model were used to evaluate a newly developed Gemcitabine-based therapy., Results: LIPH was significantly upregulated in PDAC and was related to later pathological stage and poor prognosis. LIPH downregulation in PDAC cells inhibited colony formation and proliferation. Mechanistically, LIPH triggered PI3K/AKT/HIF1A signaling via LPA/LPAR axis. LIPH also promoted glycolysis and de novo synthesis of glycerolipids by maintaining ALDOA stability in the cytosol. Xenograft models show that PDAC with high LIPH expression levels was sensitive to gemcitabine/ki16425/aldometanib therapy without causing discernible side effects., Conclusion: LIPH directly bridges PDAC cells and tumor microenvironment to facilitate aberrant aerobic glycolysis via activating LPA/LPAR axis and maintaining ALDOA stability, which provides an actionable gemcitabine-based combination therapy with limited side effects., (© 2023. The Author(s).)

Excessive intake of sugar, and particularly fructose, is closely associated with the development and progression of metabolic syndrome in humans and animal models. However, genetic disorders in fructose metabolism have very different consequences. While the deficiency of fructokinase, the first enzyme involved in fructose metabolism, is benign and somewhat desirable, missense mutations in the second enzyme, aldolase B, causes a very dramatic and sometimes lethal condition known as hereditary fructose intolerance (HFI). To date, there is no cure for HFI, and treatment is limited to avoiding fructose and sugar. Because of this, for subjects with HFI, glucose is their sole source of carbohydrates in the diet. However, clinical symptoms still occur, suggesting that either low amounts of fructose are still being consumed or, alternatively, fructose is being produced endogenously in the body. Here, we demonstrate that as a consequence of consuming high glycemic foods, the polyol pathway, a metabolic route in which fructose is produced from glucose, is activated, triggering a deleterious mechanism whereby glucose, sorbitol and alcohol induce severe liver disease and growth retardation in aldolase B knockout mice. We show that generically and pharmacologically blocking this pathway significantly improves metabolic dysfunction and thriving and increases the tolerance of aldolase B knockout mice to dietary triggers of endogenous fructose production.

The effect of ALDOA, an important regulator of tumor metabolism and immune cell function, on gastric cancer (GC) immune infiltration has not been elucidated. Hence, we explored the feasibility of using ALDOA combined with immune molecular markers as novel prognostic or therapeutic targets for GC patients. Bioinformatic analyses were initially performed in multiple databases to assess the prognostic prediction values of ALDOA expression in GC. Subsequently, both ALDOA expression and the clinicopathological characteristics of a total of 114 GC patients who underwent curative gastrectomy were collected to demonstrate the potential association between ALDOA expression and the biological behaviors of GC. Next, the expression of ALDOA and its effect on prognosis were determined at the mRNA and protein levels, respectively, using tissue microarrays and cellular experiments. Subsequently, several molecular mechanisms were revealed based on elaborate analyses, indicating that ALDOA expression was potentially involved in the progression of GC and could be considered a promising biomarker for evaluating the prognosis of GC. High ALDOA expression was frequently found in GC cells and GC tissues at the mRNA and protein levels. Based on survival analysis, the expression of ALDOA indicated comparatively poor overall survival (OS) in GC and was identified as an independent prognostic predictor of GC. Correlation analysis showed that ALDOA expression had a positive association with lymph node metastasis in GC patients. Additionally, microRNA-1179 was found to play a key role in inhibiting the expression of ALDOA in the metabolic pathways of GC cells, which might disrupt the expression of various immune molecules and be detrimental to the prognosis of GC. ALDOA should be considered a promising molecular target for evaluating the prognosis of GC, owing to its potential role in immune regulation., (© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Glucose metabolism and DNA repair are fundamental cellular processes frequently dysregulated in cancer. In this study, we define a direct role for the glycolytic Aldolase A (ALDOA) protein in DNA double-strand break (DSB) repair. ALDOA is a fructose biphosphate Aldolase that catalyses fructose-1,6-bisphosphate to glyceraldehyde 3-phosphate (G3P) and dihydroxyacetone phosphate (DHAP), during glycolysis. Here, we show that upon DNA damage induced by ionising radiation (IR), ALDOA translocates from the cytoplasm into the nucleus, where it partially co-localises with the DNA DSB marker γ-H2AX. DNA damage was shown to be elevated in ALDOA-depleted cells prior to IR and following IR the damage was repaired more slowly. Consistent with this, cells depleted of ALDOA exhibited decreased DNA DSB repair via non-homologous end-joining and homologous recombination. In support of the defective repair observed in its absence, ALDOA was found to associate with the major DSB repair effector kinases, DNA-dependent Protein Kinase (DNA-PK) and Ataxia Telangiectasia Mutated (ATM) and their autophosphorylation was decreased when ALDOA was depleted. Together, these data establish a role for an essential metabolic protein, ALDOA in DNA DSB repair and suggests that targeting ALDOA may enable the concurrent targeting of cancer metabolism and DNA repair to induce tumour cell death., (© 2023. Springer Nature Limited.)

Aldolase A (ALDOA), a crucial glycolytic enzyme, is often aberrantly expressed in various types of cancer. Although ALDOA has been reported to play additional roles beyond its conventional enzymatic role, its nonmetabolic function and underlying mechanism in cancer progression remain elusive. Here, it is shown that ALDOA promotes liver cancer growth and metastasis by accelerating mRNA translation independent of its catalytic activity. Mechanistically, ALDOA interacted with insulin- like growth factor 2 mRNA-binding protein 1 (IGF2BP1) to facilitate its binding to m 6 A-modified eIF4G mRNA, thereby increasing eIF4G protein levels and subsequently enhancing overall protein biosynthesis in cells. Importantly, administration of GalNAc-conjugated siRNA targeting ALDOA effectively slows the tumor growth of orthotopic xenografts. Collectively, these findings uncover a previously unappreciated nonmetabolic function of ALDOA in modulating mRNA translation and highlight the potential of specifically targeting ALDOA as a prospective therapeutic strategy in liver cancer., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Spermatozoa released from the testis acquire fertilizing ability by translocating thorough the epididymis. Further, accessory gland secretions ejaculated into the female reproductive tract along with spermatozoa are also required to ensure male fecundity, such as the maintenance of proper sperm count and inhibition of premature sperm capacitation in the uterus. Here, we focus on a testis-enriched gene "Aldoart2", an epididymis-enriched gene "Serpina16", and seminal vesicle-enriched genes "Aoc1l3" and "Pate14" which were thought to be important for male fertility based on the previous studies. We independently deleted almost the entire protein-coding sequence of these genes in mice using CRISPR/Cas9. There were no overt defects in the histology and the sperm morphology and motility of any knockout (KO) mice. Further, Aoc1l3 and Pate14 KO males were able to form copulatory plugs. Finally, female mice that mated with these KO males delivered pups at a comparable level with the control males. Given our data, we demonstrated that the four genes predominantly expressed in the testis, epididymis, or seminal vesicle are independently dispensable for male fertility.

In the present study, the structure of the fructose-1,6-bisphosphataldolase (FBA) gene in Mytilus galloprovincialis (Lamarck, 1819) was analyzed and its tissue specificity of expression level and activity was determined. A 1092 base pairs (bps) complete coding sequence of the FBA gene was assembled from M. galloprovincialis transcriptome. Only one gene encoding FBA (MgFBA) was identified in the M. galloprovincialis genome. The length of MgFBA was 363 amino acids with a molecular mass of 39.7 kDa. According to the amino acid residues, the detected MgFBA gene is a type I aldolase. The FBA gene in M. galloprovincialis had 7 exons; the maximum intron length was about 2.5 kbps. Intraspecific nucleotide diversity (15 mutations) between MgFBAs from the Mediterranean mussels and the Black Sea mussels (present study) was detected. All mutations were synonymous. Tissue specificity in FBA expression level and activity was established. No direct correlation between these functions was found. The highest level of FBA gene expression is found in muscle tissue. According to the phylogenetic analyses, FBA gene in invertebrates could be considered the ancestral gene of muscle type aldolase, which may explain the character of tissue-specific expression., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to declare that they are relevant to the content of this article. All applicable institutional, national and international guidelines for the care and use of animals were followed., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Colorectal cancer (CRC) is a serious threat to human health, and its underlying mechanisms remain to be further explored. Aldolase A (ALDOA) has received increasing attention for its reported association with multiple cancers, but the role and mechanisms of ALDOA in CRC are still unclear. In the current study, high expression levels and enzymatic activity of ALDOA were detected in CRC tissues and cell lines, indicating the clinical significance of ALDOA in human CRC. In addition, silencing ALDOA significantly impaired the proliferation and metastasis of CRC cells in vitro and in vivo . Mechanistically, immunoprecipitation assays and mass spectrometry analysis identified the binding protein COPS6 of ALDOA. Furthermore, the promoting effects of upregulated ALDOA on CRC cell proliferation and metastasis were inhibited by COPS6 depletion, demonstrating COPS6 was required for ALDOA in mediating CRC progress. Moreover, the epithelial-mesenchymal transition (EMT) program and MAPK signaling pathway were found to be activated by ALDOA overexpression as well. In summary, our findings suggested that ALDOA facilitated the proliferation and metastasis of CRC by binding and regulating COPS6, inducing EMT, and activating the mitogen-activated protein kinase (MAPK) signaling pathway. The present study provided evidence for ALDOA as a promising potential biomarker for CRC., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this article., (Copyright © 2023 Ya Lu et al.)

LncRNA PSMA3-AS1 functions as an oncogene in several cancers, including ovarian cancer, lung cancer, and colorectal cancer. However, its role in gastric cancer (GC) progression remains unclear. In this study, the levels of PSMA3-AS1, miR-329-3p, and aldolase A (ALDOA) in 20 paired human GC tissues and adjacent nontumorous tissues were measured by real-time PCR. GC cells were transfected with recombinant plasmid carrying full-length PSMA3-AS1 or shRNA targeting PSMA3-AS1. The stable transfectants were selected by G418. Then, the effects of PSMA3-AS1 knockdown or overexpression on GC progression in vitro and in vivo were evaluated. The results showed that PSMA3-AS1 was highly expressed in human GC tissues. Stable knockdown of PSMA3-AS1 significantly restrained proliferation/migration/invasion, enhanced cell apoptosis, and induced oxidative stress in vitro. Tumor growth and matrix metalloproteinase expression in tumor tissues were markedly inhibited, while oxidative stress was enhanced in nude mice after stable PSMA3-AS1 knockdown. Additionally, PSMA3-AS1 negatively regulated miR-329-3p while positively regulated ALDOA expression. MiR-329-3p directly targeted ALDOA-3'UTR. Interestingly, miR-329-3p knockdown or ALDOA overexpression partially attenuated the tumor-suppressive effects of PSMA3-AS1 knockdown. Conversely, PSMA3-AS1 overexpression exhibited the opposite effects. PSMA3-AS1 promoted GC progression by regulating the miR-329-3p/ALDOA axis. PSMA3-AS1 might serve as a promising and effective target for GC treatment., (© 2023. The Author(s).)

Aldolase B (ALDOB), a glycolytic enzyme, is uniformly depleted in clear cell renal cell carcinoma (ccRCC) tissues. We previously showed that ALDOB inhibited proliferation through a mechanism independent of its enzymatic activity in ccRCC, but the mechanism was not unequivocally identified. We showed that the corepressor C-terminal-binding protein 2 (CtBP2) is a novel ALDOB-interacting protein in ccRCC. The CtBP2-to-ALDOB expression ratio in clinical samples was correlated with the expression of CtBP2 target genes and was associated with shorter survival. ALDOB inhibited CtBP2-mediated repression of multiple cell cycle inhibitor, proapoptotic, and epithelial marker genes. Furthermore, ALDOB overexpression decreased the proliferation and migration of ccRCC cells in an ALDOB-CtBP2 interaction-dependent manner. Mechanistically, our findings showed that ALDOB recruited acireductone dioxygenase 1, which catalyzes the synthesis of an endogenous inhibitor of CtBP2, 4-methylthio 2-oxobutyric acid. ALDOB functions as a scaffold to bring acireductone dioxygenase and CtBP2 in close proximity to potentiate acireductone dioxygenase-mediated inhibition of CtBP2, and this scaffolding effect was independent of ALDOB enzymatic activity. Moreover, increased ALDOB expression inhibited tumor growth in a xenograft model and decreased lung metastasis in vivo. Our findings reveal that ALDOB is a negative regulator of CtBP2 and inhibits tumor growth and metastasis in ccRCC., (© 2023. Higher Education Press.)

Background: Ticks are obligate hematophagous ectoparasites that transmit a variety of pathogens to humans, wildlife and domestic animals. Vaccination is an effective and environmentally friendly method for tick control. Fructose-1,6-bisphosphate aldolase (FBA) is an important glycometabolism enzyme that is a candidate vaccine against parasites. However, the immune protection of FBA in ticks is unclear. METHODS AND RESULTS: The 1092-bp open reading frame (ORF) of FBA from Haemaphysalis longicornis (HlFBA), encoding a 363-amino acid protein, was cloned using PCR methodology. The prokaryotic expression vector pET32a(+)-HlFBA was constructed and transformed into cells of Escherichia coli BL21(DE3) strain for protein expression. The recombinant HlFBA protein (rHlFBA) was purified by affinity chromatography, and the western blot results suggested that the rHlFBA protein was immunogenic., Results: Results of the enzyme-linked immunosorbent assay showed that rabbits immunized with rHlFBA produced a humoral immune response specific to rHlFBA. A tick infestation trial indicated that, compared to the ticks in the histidine-tagged thioredoxin (Trx) group, the engorged tick weight and oviposition of female ticks and egg hatching rate of those in the rHlFBA group was reduced by 22.6%, 45.6% and 24.1%, respectively. Based on the cumulative effect of the these three parameters, the overall immune efficacy of rHlFBA was estimated to be 68.4%., Conclusions: FBA is a candidate anti-tick vaccine that can significantly reduce the engorged tick weight, oviposition, and egg hatching rate. The use of enzymes involved in glucose metabolism is a new strategy in the development of anti-tick vaccines., (© 2023. The Author(s).)

Colorectal cancer (CRC) is an aggressive tumor, whose development is considered to be modulated by certain long non‑coding RNAs (lncRNAs). Therefore, the aim of the present study was to investigate the regulatory mechanism of lncRNA NONHSAG028908.3 on CRC. Data from The Cancer Genome Atlas (TCGA) database revealed that NONHSAG028908.3 was increased in CRC tissues compared with normal tissues (P<0.001). The results of reverse transcription‑quantitative PCR indicated that NONHSAG028908.3 was upregulated in four types of CRC cells compared with that in NCM460, a normal colorectal cell line. MTT, BrdU, and flow cytometric assays were applied to evaluate CRC cell growth. The migratory and invasive abilities of CRC cells were detected using wound healing and Transwell assays. Silencing of NONHSAG028908.3 inhibited proliferation, migration, and invasion of CRC cells. A dual‑luciferase reporter assay demonstrated that NONHSAG028908.3 served as a sponge to combine with microRNA (miR)‑34a‑5p. MiR‑34a‑5p suppressed the aggressiveness of CRC cells. The effects induced by NONHSAG028908.3 knockdown were partly reversed by inhibition of miR‑34a‑5p. Furthermore, miR‑34a‑5p, a target of NONHSAG028908.3, modulated aldolase, fructose‑bisphosphate A (ALDOA) expression in a negative feedback manner. Suppression of NONHSAG028908.3 notably decreased ALDOA expression, which was rescued via silencing of miR‑34a‑5p. Moreover, suppression of ALDOA revealed the inhibitory action on CRC cell growth and migration. In summary, the data of the present study indicate that NONHSAG028908.3 may positively regulate ALDOA via sponging miR‑34a‑5p, thereby promoting malignant activities in CRC.

Aldolase B (ALDOB) gene is essential for the process of glycolysis and differentially expressed in cancers. The aims of this study were to explore the potential role of ALDOB in pan-cancer, in order to deepen the research on the pathological mechanism of cancer. Hence, we used several online tools (TIMER2, GEPIA2, UALCAN, cBioPortal, and MXPRESS) and R language to identify the correlation between the ALDOB expression and survival analysis, genetic alteration, DNA methylation, and immune cell infiltration based on The Cancer Genome Atlas project. The results showed that ALDOB was lowly expressed in pan-cancer. Survival analysis revealed that low expression of ALDOB was markedly related with poor clinical prognosis, while the genetic alteration within ALDOB changed along with the difference of overall survival (OS) and disease-free survival (DFS) prognosis in several cancers. A possible relationship between DNA methylation and ALDOB expression for several tumors was found. Besides, ALDOB expression was confirmed to be associated with tumor immune cell infiltration, especially in breast invasive carcinoma (BRCA), esophageal carcinoma (ESCA), and testicular germ cell tumors (TGCT) cases. Further, the enrichment analysis demonstrated that metabolic pathway was closely related to ALDOB expression. Our results provide a comprehensive pan-cancer analysis and suggest ALDOB could act as a promising tumor predictive biomarker for human cancer., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2023 the Author(s). Published by Wolters Kluwer Health, Inc.)

The preferred carbon source of Staphylococcus aureus and many other pathogens is glucose, and its consumption is critical during infection. However, glucose utilization increases the cellular demand for manganese, a nutrient sequestered by the host as a defense against invading pathogens. Therefore, bacteria must balance glucose metabolism with the increasing demand that metal-dependent processes, such as glycolysis, impose upon the cell. A critical regulator that enables S. aureus to resist nutritional immunity is the ArlRS two-component system. This work revealed that ArlRS regulates the expression of FdaB, a metal-independent fructose 1,6-bisphosphate aldolase. Further investigation revealed that when S. aureus is metal-starved by the host, FdaB functionally replaces the metal-dependent isozyme FbaA, thereby allowing S. aureus to resist host-imposed metal starvation in culture. Although metal-dependent aldolases are canonically zinc-dependent, this work uncovered that FbaA requires manganese for activity and that FdaB protects S. aureus from manganese starvation. Both FbaA and FdaB contribute to the ability of S. aureus to cause invasive disease in wild-type mice. However, the virulence defect of a strain lacking FdaB was reversed in calprotectin-deficient mice, which have defects in manganese sequestration, indicating that this isozyme contributes to the ability of this pathogen to overcome manganese limitation during infection. Cumulatively, these observations suggest that the expression of the metal-independent aldolase FdaB allows S. aureus to alleviate the increased demand for manganese that glucose consumption imposes, and highlights the cofactor flexibility of even established metalloenzyme families. IMPORTANCE Staphylococcus aureus and other pathogens consume glucose during infection. Glucose utilization increases the demand for transition metals, such as manganese, a nutrient that the host limits as a defense mechanism against invading pathogens. Therefore, pathogenic bacteria must balance glucose and manganese requirements during infection. The two-component system ArlRS is an important regulator that allows S. aureus to adapt to both glucose and manganese starvation. Among the genes regulated by ArlRS is the metal-independent fructose 1,6-bisphosphate aldolase fdaB , which functionally substitutes for the metal-dependent isoenzyme FbaA and enables S. aureus to survive host-imposed manganese starvation. Unexpectedly, and differing from most characterized metal-dependent aldolases, FbaA requires manganese for activity. Cumulatively, these findings reveal a new mechanism for overcoming nutritional immunity as well as the cofactor plasticity of even well-characterized metalloenzyme families.

The raw material cost of lactic acid fermentation accounts for the main part of the production cost, and this necessitates the exploration of the efficient use of cheap raw materials in lactic acid production. We compared the outcomes of the homologous expressions of xylose transporters (xylFGH, xylE, araE, and xylT), 6-phosphofructokinase (pfkA), fructose-bisphosphate aldolase (fbaA), and their co-expression in Lactococcus lactis IO-1 on lactic acid production using xylose as the raw material. We found that the production rate of lactic acid on xylose fermentation by L. lactis IO-1 overexpressing fbaA was the highest (14.42%). Among the xylose transporters investigated, XylT had the strongest xylose transport capacity in L. lactis IO-1, with an increase in the lactic acid production rate by 10.38%. The genes near the overexpression of fbaA or xylT in the metabolic pathway were more upregulated than the distant genes. The co-expression of fbaA and xylT increased the production rate of lactic acid by 27.84% on xylose fermentation by L. lactis IO-1. This work presents a novel strategy for the simultaneous enhancement of the expression of important genes at the beginning and midway of the xylose metabolic pathway of L. lactis IO-1, which could greatly improve the target production., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Available therapeutics for visceral leishmaniasis (VL), a deadly parasitic infection, are usually associated with inadequate efficacy and adverse aftereffects. Further, the primary site of Leishmania parasite are host macrophages resulting in compromised immunity; ensuing marked T-cell immunosuppression. Such settings emphasize the exploration of chemo-immunotherapeutic strategies for improvising the infected person's immune status with better resolution of infection., Methods: Present work employs the immunization of Leishmania-infected hamsters with Leishmania-derived recombinant aldolase (rLdAld) and enolase (rLdEno) proteins in consort with the sub-optimal dose of Ambisome (2.5 mg/kg). After the completion of immunization, hamsters were sacrificed on day 60 and 90 post infection and different organ samples were collected to perform immunological assay for evaluating the therapeutic efficacy and modulation in protective cellular immune responses., Results: Combining these proteins, particularly rLdAld with Ambisome (2.5 mg/kg), has significantly reduced the parasitic load (∼80%) with remarkable enhancement in DTH and lymphoproliferative responses compared to the infected control and only Ambisome treated groups. Moreover, cytokine levels at RNA and protein levels were noticed to be inclined towards Th-1 phenotype through up-regulation of IFN-γ and TNF-α with significant down-regulation in IL-10 and TGF-β expression, an indication towards the generation of protective immunity against experimental VL., Conclusion: Our experimental findings demonstrated that the chemo-immunotherapeutic approach could be an effective way of controlling human VL infection at minimal dosages of antileishmanial with reduced side-effects and propensity of drug resistance emergence., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2022. Published by Elsevier B.V.)

NahE is a hydratase-aldolase that converts o-substituted trans-benzylidenepyruvates (H, OH, or CO 2 - ) to benzaldehyde, salicylaldehyde, or 2-carboxybenzaldehyde, respectively, and pyruvate. The enzyme is in a bacterial degradative pathway for naphthalene, which is a toxic and persistent environmental contaminant. Sequence, crystallographic, and mutagenic analysis identified the enzyme as a member of the N-acetylneuraminate lyase (NAL) subgroup in the aldolase superfamily. As such, it has a conserved lysine (Lys183) and tyrosine (Tyr155), for Schiff base formation, as well as a GXXGE motif for binding of the pyruvoyl carboxylate group. A crystal structure of the selenomethionine derivative of NahE shows these active site elements along with nearby residues that might be involved in the mechanism and/or specificity. Mutations of five active site amino acids (Thr65, Trp128, Tyr155, Asn157, and Asn281) were constructed and kinetic parameters measured in order to assess the effect(s) on catalysis. The results show that the two Trp128 mutants (Phe and Tyr) have the least effect on catalysis, whereas amino acids with bulky side chains at Thr65 (Val) and Asn281 (Leu) have the greatest effect. Changing Tyr155 to Phe and Asn157 to Ala also hinders catalysis, and the effects fall in between these extremes. These observations are put into a structural context using a crystal structure of the Schiff base of the reaction intermediate. Trapping experiments with substrate, Na(CN)BH 3 , and wild type enzyme and selected mutants mostly paralleled the kinetic analysis, and identified two salicylaldehyde-modified lysines: the active site lysine (Lys183) and one outside the active site (Lys279). The latter could be responsible for the observed inhibition of NahE by salicylaldehyde. Together, the results provide new insights into the NahE-catalyzed reaction., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Aldolase A (ALDOA), an important metabolic enzyme in the glycolytic pathway, plays an important role in regulating tumour metabolism. In this study, we investigated the expression pattern of ALDOA in hepatocellular carcinoma (HCC) and its biological role in tumour progression. Bioinformatics analysis, western blot (WB) and RT-qPCR were performed to detect the relative expression of ALDOA in HCC tissues and cell lines. A loss-of-function approach was used to investigate the biological function of ALDOA. The role of ALDOA on glycolysis was assessed by WB, glucose and lactate assay kits and a nude mouse xenograft model. Luciferase reporter experiment, chromatin immunoprecipitation and WB were performed to elucidate the underlying molecular. The expression level of ALODA was up-regulated in HCC tissues and cell lines. High ALDOA levels were associated with poorer patient overall survival. Mechanistic studies suggest that ALDOA is a direct target of miR-34a-5p, which can inhibit glycolysis in hepatocellular carcinoma cells by targeting the 3'UTR of ALDOA. PINK1 antisense RNA (PINK1-AS) competitively sponged miR-34a-5p to increase ALDOA expression by antagonizing miR-34a-5p-mediated ALDOA inhibition. MKL-1 acted as a transcription factor to promote the expression of PINK1-AS and ALDOA, thus promoting the deterioration of HCC cells. This study shows that high expression of ALDOA contributes to the development and poor prognosis of hepatocellular carcinoma and will be a target and potential prognostic biomarker for the treatment of HCC., (© 2022. The Author(s).)

Eimeria tenella (E. tenella) is a protozoal parasite that can cause severe cecal lesions and death in chickens, seriously harming the chicken industry. Conventional control strategies mainly rely on anticoccidial drugs. However, the emerging problems of anticoccidial resistance and drug residues necessitate exploring potential drug targets for developing new anticoccidial drugs. Fructose-1,6-bisphosphate aldolase (ALD) is an essential enzyme for parasite energy metabolism that has been considered a potential drug target. In this study, we analyzed the molecular and biochemical properties of E. tenella ALD2 (EtALD2). EtALD2 mRNA expression was highest in second-generation merozoites, whereas the protein level was highest in unsporulated oocysts. Indirect immunofluorescence showed that EtALD2 was mainly distributed in sporozoite' cytoplasm. The natural product inhibitor (morin) was screened by computer-aided drug screening. Enzyme kinetic and inhibition kinetic assays showed that morin had a good inhibitory effect on EtALD2 activity (IC 50  = 10.37 μM, Ki = 48.97 μM). In vitro inhibition assay demonstrated that morin had an inhibitory effect on E. tenella development, with an IC 50 value of 3.98 μM and drug selection index of 177.49. In vivo, morin significantly improved cecal lesions (p < 0.05) and reduced oocyst excretion (p < 0.05) in E. tenella-infected chickens compared with the untreated group. The anticoccidial index of the group receiving 450 mg morin per kg feed was 162, showing a good anticoccidial effect. These findings suggest that EtALD2 could be a novel drug target for E. tenella treatment, and morin should be further evaluated as a therapeutic candidate for chicken coccidiosis., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interests regarding the publication of this paper., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Nitric oxide as a signaling molecule exerts cytotoxicity known as nitrosative stress at its excess concentrations. In the yeast Saccharomyces cerevisiae, the cellular responses to nitrosative stress and their molecular mechanisms are not fully understood. Here, focusing on the posttranslational modifications that are associated with nitrosative stress response, we show that nitrosative stress increased the protein S-glutathionylation level in yeast cells. Our proteomic and immunochemical analyses demonstrated that the fructose-1,6-bisphosphate aldolase Fba1 underwent S-glutathionylation at Cys112 in response to nitrosative stress. The enzyme assay using a recombinant Fba1 demonstrated that S-glutathionylation at Cys112 inhibited the Fba1 activity. Moreover, we revealed that the cytosolic glutaredoxin Grx1 reduced S-glutathionylation of Fba1 and then recovered its activity. The intracellular contents of fructose-1,6-bisphosphate and 6-phosphogluconate, which are a substrate of Fba1 and an intermediate of the pentose phosphate pathway (PPP), respectively, were increased in response to nitrosative stress, suggesting that the metabolic flow was switched from glycolysis to PPP. The cellular level of NADPH, which is produced in PPP and functions as a reducing force for nitric oxide detoxifying enzymes, was also elevated under nitrosative stress conditions, but this increase was canceled by the amino acid substitution of Cys112 to Ser in Fba1. Furthermore, the viability of yeast cells expressing Cys112Ser-Fba1 was significantly lower than that of the wild-type cells under nitrosative stress conditions. These results indicate that the inhibition of Fba1 by its S-glutathionylation changes metabolism from glycolysis to PPP to increase NADPH production, leading to nitrosative stress tolerance in yeast cells., Competing Interests: Declaration of competing interest The authors declare no conflict of interests., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Fructose-1,6-bisphosphate aldolase (EC 4.1.2.13) is a highly conserved enzyme that is involved in glycolysis and gluconeogenesis. In this study, we cloned the fructose-1,6-bisphosphate aldolase gene from Euphausia superba (EsFBA). The full-length cDNA sequence of EsFBA is 1098 bp long and encodes a 365-amino-acid protein. The fructose-1,6-bisphosphate aldolase gene was expressed in Escherichia coli ( E. coli ). A highly purified protein was obtained using HisTrap HP affinity chromatography and size-exclusion chromatography. The predicted three-dimensional structure of EsFBA showed a 65.66% homology with human aldolase, whereas it had the highest homology (84.38%) with the FBA of Penaeus vannamei . Recombinant EsFBA had the highest activity at 45 °C and pH 7.0 in phosphate buffer. By examining the activity of metal ions and EDTA, we found that the effect of metal ions and EDTA on EsFBA's enzyme activity was not significant, while the presence of borohydride severely reduced the enzymatic activity; thus, EsFBA was confirmed to be a class I aldolase. Furthermore, targeted mutations at positions 34, 147, 188, and 230 confirmed that they are key amino acid residues for EsFBA.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a baculovirus that infects silkworms, and its interaction with silkworm has been considered an important model in the field of insect virology. Accumulating evidence indicates that most viruses promote glycolytic metabolism in host cells to favor infection. However, similar reports are lacking in insects, especially in the area of post-translational modifications of proteins. In this study, we found that BmNPV infection induced the acetylation of fructose-bisphosphate aldolase (ALDO) on lysine 42 (K42) to promote its enzyme activity. To explore the underlying mechanisms, site-directed mutagenesis of deacetylated mimic (K/R) was performed. The results demonstrated that K42 acetylation promoted viral proliferation by exacerbating the glycolytic flux induced by BmNPV infection, which resulted in increased ATP, glucose uptake and lactate accumulation. Inhibiting glycolysis with 2-deoxygucose (2DG) revealed that glycolysis was essential for optimal BmNPV infection. Finally, we showed that BmNPV-infected cells enhanced the transcription of glycolysis-related genes, including Glut1, Hk2 and Ldh. In parallel, K42 acetylation of ALDO also promoted the expression of these genes. Therefore, acetylation of ALDO could be considered a regulator of BmNPV-induced glycolysis. These finding provide insights into the interaction between silkworm and BmNPV., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

To successfully design expression systems for industrial biotechnology and biopharmaceutical applications; plasmid stability, efficient synthesis of the desired product and the use of selection markers acceptable to regulatory bodies are of utmost importance. In this work we demonstrate the application of a set of IPTG-inducible protein expression systems -- harboring different features namely, antibiotic vs auxotrophy marker; two-plasmids vs single plasmid expression system; expression levels of the repressor protein (LacI) and the auxotrophic marker (glyA) -- in high-cell density cultures to evaluate their suitability in bioprocess conditions that resemble industrial settings. Results revealed that the first generation of engineered strain showed a 50% reduction in the production of the model recombinant protein fuculose-1-phosphate aldolase (FucA) compared to the reference system from QIAGEN. The over-transcription of glyA was found to be a major factor responsible for the metabolic burden. The second- and third-generation of expression systems presented an increase in FucA production and advantageous features. In particular, the third-generation expression system is antibiotic-free, autotrophy-selection based and single-plasmid and, is capable to produce FucA at similar levels compared to the original commercial expression system. These new tools open new avenues for high-yield and robust expression of recombinant proteins in E. coli., (© The Author(s) 2022. Published by Oxford University Press on behalf of Society of Industrial Microbiology and Biotechnology.)

Aldolase A (ALDOA) is an enzyme that plays an important role in glycolysis and gluconeogenesis, which is closely related to tumor metabolism. In this study, the overall roles of ALDOA in pan-cancer have been investigated from several aspects using databases and online analysis tools. Using the ONCOMINE database, the expression of ALDOA in various cancers was analyzed. The prognostic role of ALDOA was explored by PrognoScan, GEPIA, and Kaplan-Meier Plotter. The immune-related role of ALDOA and its downstream substrates was decided by TIMER, cBioPortal and String. Our data indicate that ALDOA expression level in lung adenocarcinoma, liver hepatocellular carcinoma, head and neck squamous cell carcinoma is higher than that in normal tissues. Increased expression of ALDOA often indicates a poor prognosis for patients. The correlation between ALDOA and immune infiltration among different tumors is very different. We also investigate the relationship between ALDOA and its upstream/downstream proteins. Our results showed that ALDOA could be used as a biomarker for the tumor prognosis, and could be correlated with the infiltrating levels of macrophages, CD4+ T cells and CD8+ T cells., (© 2022. The Author(s).)

Metabolic reprogramming and elevated glycolysis levels are associated with tumor progression. However, despite cancer cells selectively inhibiting or expressing certain metabolic enzymes, it is unclear whether differences in gene profiles influence patient outcomes. Therefore, identifying the differences in enzyme action may facilitate discovery of gene ontology variations to characterize tumors. Fructose-1,6-bisphosphate (F-1,6-BP) is an important intermediate in glucose metabolism, particularly in cancer. Gluconeogenesis and glycolysis require fructose-1,6-bisphosphonates 1 (FBP1) and fructose-bisphosphate aldolase A (ALDOA), which participate in F-1,6-BP conversion. Increased expression of ALDOA and decreased expression of FBP1 are associated with the progression of various forms of cancer in humans. However, the exact molecular mechanism by which ALDOA and FBP1 are involved in the switching of F-1,6-BP is not yet known. As a result of their pancancer pattern, the relationship between ALDOA and FBP1 in patient prognosis is reversed, particularly in lung adenocarcinoma (LUAD) and liver hepatocellular carcinoma (LIHC). Using The Cancer Genome Atlas (TCGA), we observed that FBP1 expression was low in patients with LUAD and LIHC tumors, which was distinct from ALDOA. A similar trend was observed in the analysis of Cancer Cell Line Encyclopedia (CCLE) datasets. By dissecting downstream networks and possible upstream regulators, using ALDOA and FBP1 as the core, we identified common signatures and interaction events regulated by ALDOA and FBP1. Notably, the identified effectors dominated by ALDOA or FBP1 were distributed in opposite patterns and can be considered independent prognostic indicators for patients with LUAD and LIHC. Therefore, uncovering the effectors between ALDOA and FBP1 will lead to novel therapeutic strategies for cancer patients.

Colorectal cancer (CRC) is a leading cause of cancer-related death worldwide. The spheroid colony formation assay is a useful method to identify cancer stem cells (CSCs). Using the DLD-1 and WiDr CRC cell lines, we performed microarray analyses of spheroid body-forming and parental cells and demonstrated that aldolase, fructose-bisphosphate C (ALDOC) was overexpressed in the spheroid body-forming cells of both lines. Cells transfected with small interfering RNA against ALDOC demonstrated lower proliferation, migration, and invasion compared with negative control cells. Both the number and size of spheres produced by the CRC cells were significantly reduced by ALDOC knockdown. Additionally, inhibition of ALDOC reduced lactate production. Immunohistochemistry was used to analyze ALDOC protein expression in tissues from 135 CRC patients and revealed that 66 (49%) cases were positive for ALDOC. The ALDOC-positive cases were associated with higher T and M grades and, as determined by Kaplan-Meier analysis, a poorer prognosis. Univariate and multivariate analyses indicated that ALDOC expression was an independent prognostic factor for CRC patients. Furthermore, ALDOC expression was associated with CD44 expression. These results suggest that ALDOC contributes to CRC progression and plays an important role in CSCs derived from CRC., (© 2022 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.)

Some protein positions play special roles in determining the magnitude of protein function: at such "rheostat" positions, varied amino acid substitutions give rise to a continuum of functional outcomes, from wild type (or enhanced), to intermediate, to loss of function. This observed range raises interesting questions about the biophysical bases by which changes at single positions have such varied outcomes. Here, we assessed variants at position 98 in human aldolase A ("I98X"). Despite being ~17 Å from the active site and far from subunit interfaces, substitutions at position 98 have rheostatic contributions to the apparent cooperativity (n H ) associated with fructose-1,6-bisphosphate substrate binding and moderately affected binding affinity. Next, we crystallized representative I98X variants to assess structural consequences. Residues smaller than the native isoleucine (cysteine and serine) were readily accommodated, and the larger phenylalanine caused only a slight separation of the two parallel helixes. However, the diffraction quality was reduced for I98F, and further reduced for I98Y. Intriguingly, the resolutions of the I98X structures correlated with their n H values. We propose that substitution effects on both n H and crystal lattice disruption arise from changes in the population of aldolase A conformations in solution. In combination with results computed for rheostat positions in other proteins, the results from this study suggest that rheostat positions accommodate a wide range of side chains and that structural consequences manifest as shifted ensemble populations and/or dynamics changes., (© 2021 The Protein Society.)

Background: Glycolysis dysfunction is an important pathogenesis of podocyte injury in diabetic kidney disease (DKD). Foot process fusion of podocytes and increased albuminuria are markers of early DKD. Moreover, cytoskeletal remodeling has been found to be involved in the foot process fusion of podocytes. However, the connections between cytoskeletal remodeling and alterations of glycolysis in podocytes in DKD have not been clarified., Methods: mRNA sequencing of glomeruli obtained from db/db and db/m mice with albuminuria was performed to analyze the expression profiling of genes in glucose metabolism. Expressions of phosphofructokinase platelet type (PFKP) in the glomeruli of DKD patients were detected. Clotrimazole (CTZ) was used to explore the renal effects of PFKP inhibition in diabetic mice. Using Pfkp siRNA or recombinant plasmid to manipulate PFKP expression, the effects of PFKP on high glucose (HG) induced podocyte damage were assessed in vitro . The levels of fructose-1,6-bisphosphate (FBP) were measured. Targeted metabolomics was performed to observe the alterations of the metabolites in glucose metabolism after HG stimulation. Furthermore, aldolase type b (Aldob) siRNA or recombinant plasmid were applied to evaluate the influence of FBP level alteration on podocytes. FBP was directly added to podocyte culture media. Db/db mice were treated with FBP to investigate its effects on their kidney., Results: mRNA sequencing showed that glycolysis enzyme genes were altered, characterized by upregulation of upstream genes ( Hk1, and Pfkp ) and down-regulation of downstream genes of glycolysis ( Pkm, and Ldha ). Moreover, the expression of PFKP was increased in glomeruli of DKD patients. The CTZ group presented more severe renal damage. In vitro , the Pfkp siRNA group and ALDOB overexpression group showed much more induced cytoskeletal remodeling in podocytes, while overexpression of PFKP and suppression of ALDOB in vitro rescued podocytes from cytoskeletal remodeling through regulation of FBP levels and inhibition of the RhoA/ROCK1 pathway. Furthermore, targeted metabolomics showed FBP level was significantly increased in HG group compared with the control group. Exogenous FBP addition reduced podocyte cytoskeletal remodeling and renal damage of db/db mice., Conclusions: These findings provide evidence that PFKP may be a potential target for podocyte injury in DN and provide a rationale for applying podocyte glycolysis enhancing agents in patients with DKD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zhang, Liang, Luo, Hu, Yang, Hu, Chen, Zhu, Feng, Zhu, Chi and Ding.)

The milk yeast Kluyveromyces lactis degrades glucose through glycolysis and the pentose phosphate pathway and follows a mainly respiratory metabolism. Here, we investigated the role of two reactions which are required for the final steps of glucose degradation from both pathways, as well as for gluconeogenesis, namely fructose-1,6-bisphosphate aldolase (FBA) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In silico analyses identified one gene encoding the former ( KlFBA1 ), and three genes encoding isoforms of the latter ( KlTDH1 , KlTDH2 , KlGDP1 ). Phenotypic analyses were performed by deleting the genes from the haploid K. lactis genome. While Klfba1 deletions lacked detectable FBA activity, they still grew poorly on glucose. To investigate the in vivo importance of the GAPDH isoforms, different mutant combinations were analyzed for their growth behavior and enzymatic activity. KlTdh2 represented the major glycolytic GAPDH isoform, as its lack caused a slower growth on glucose. Cells lacking both KlTdh1 and KlTdh2 failed to grow on glucose but were still able to use ethanol as sole carbon sources, indicating that KlGdp1 is sufficient to promote gluconeogenesis. Life-cell fluorescence microscopy revealed that KlTdh2 accumulated in the nucleus upon exposure to oxidative stress, suggesting a moonlighting function of this isoform in the regulation of gene expression. Heterologous complementation of the Klfba1 deletion by the human ALDOA gene renders K. lactis a promising host for heterologous expression of human disease alleles and/or a screening system for specific drugs.

Hereditary fructose intolerance (HFI) is an inborn error of fructose metabolism of autosomal recessive inheritance caused by pathogenic variants in the ALDOB gene that lead to aldolase B deficiency in the liver, kidneys, and intestine. Patients manifest symptoms, such as ketotic hypoglycemia, vomiting, nausea, in addition to hepatomegaly and other liver and kidney dysfunctions. The treatment consists of a fructose-restricted diet, which results in a good prognosis. To analyze the distribution of ALDOB variants described in patients and to estimate the prevalence of HFI based on carrier frequency in the gnomAD database, a systematic review was conducted to assess ALDOB gene variants among patients with HFI. The prevalence of HFI was estimated from the carrier frequency of variants described in patients, as well as rare variants predicted as pathogenic by in silico tools. The p.(Ala150Pro) and p.(Ala175Asp) variants are the most frequent and are distributed worldwide. However, these variants have particular distribution patterns in Europe. The analysis of the prevalence of HFI showed that the inclusion of rare alleles predicted as pathogenic is a more informative approach for populations with few patients. The data show that HFI has a wide distribution and an estimated prevalence of ~1:10,000., (© 2021 Wiley Periodicals LLC.)

Background and Aims: Insulin receptor (IR) transduces cell surface signal through phosphoinositide 3-kinase (PI3K)-AKT pathways or translocates to the nucleus and binds to the promoters to regulate genes associated with insulin actions, including de novo lipogenesis (DNL). Chronic activation of IR signaling drives malignant transformation, but the underlying mechanisms remain poorly defined. Down-regulation of fructose-1,6-bisphosphate aldolase (ALDO) B in hepatocellular carcinoma (HCC) is correlated with poor prognosis. We aim to study whether and how ALDOB is involved in IR signaling in HCC., Approach and Results: Global or liver-specific ALDOB knockout (L-ALDOB -/- ) mice were used in N-diethylnitrosamine (DEN)-induced HCC models, whereas restoration of ALDOB expression was achieved in L-ALDOB -/- mice by adeno-associated virus (AAV). 13 C 6 -glucose was employed in metabolic flux analysis to track the de novo fatty acid synthesis from glucose, and nontargeted lipidomics and targeted fatty acid analysis using mass spectrometry were performed. We found that ALDOB physically interacts with IR and attenuates IR signaling through down-regulating PI3K-AKT pathways and suppressing IR nuclear translocation. ALDOB depletion or disruption of IR/ALDOB interaction in ALDOB mutants promotes DNL and tumorigenesis, which is significantly attenuated with ALDOB restoration in L-ALDOB -/- mice. Notably, attenuated IR/ALDOB interaction in ALDOB-R46A mutant exhibits more significant tumorigenesis than releasing ALDOB/AKT interaction in ALDOB-R43A, whereas knockdown IR sufficiently diminishes tumor-promoting effects in both mutants. Furthermore, inhibiting phosphorylated AKT or fatty acid synthase significantly attenuates HCC in L-ALDOB -/- mice. Consistently, ALDOB down-regulation is correlated with up-regulation of IR signaling and DNL in human HCC tumor tissues., Conclusions: Our study reports a mechanism by which loss of ALDOB activates IR signaling primarily through releasing IR/ALDOB interaction to promote DNL and HCC, highlighting a potential therapeutic strategy in HCC., (© 2021 by the American Association for the Study of Liver Diseases.)

Cardiovascular diseases (CVD), with myocardial infarction (MI) being one of the crucial components, wreak havoc in developed countries. Advanced imaging technologies are required to obtain quick and widely available diagnostic data. This paper describes a multimodal approach to in vivo perfusion imaging using the novel SYN1 tracer based on the fluorine-18 isotope. The NOD-SCID mice were injected intravenously with SYN1 or [ 18 F] fluorodeoxyglucose ([ 18 F]-FDG) radiotracers after induction of the MI. In all studies, the positron emission tomography-computed tomography (PET/CT) technique was used. To obtain hemodynamic data, mice were subjected to magnetic resonance imaging (MRI). Finally, the biodistribution of the SYN1 compound was performed using Wistar rat model. SYN1 showed normal accumulation in mouse and rat hearts, and MI hearts correctly indicated impaired cardiac segments when compared to [ 18 F]-FDG uptake. In vivo PET/CT and MRI studies showed statistical convergence in terms of the size of the necrotic zone and cardiac function. This was further supported with RNAseq molecular analyses to correlate the candidate function genes' expression, with Serpinb1c , Tnc and Nupr1 , with Trem2 and Aldolase B functional correlations showing statistical significance in both SYN1 and [ 18 F]-FDG. Our manuscript presents a new fluorine-18-based perfusion radiotracer for PET/CT imaging that may have importance in clinical applications. Future research should focus on confirmation of the data elucidated here to prepare SYN1 for first-in-human trials.