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The agricultural pest Drosophila suzukii exhibits a strong preference for feeding on fresh fruits, demonstrating high adaptability to sugary environments. Meanwhile, high sugar levels stimulate insulin secretion, thereby regulating the steady state of sugar metabolism. Understanding the mechanisms related to sugar metabolism in D. suzukii is crucial due to its adaptation to these specific environmental conditions. The insulin signaling pathway is an evolutionarily conserved phosphorylation cascade with significant roles in development and metabolism. We observed that the activation of the insulin signaling pathway inhibited FoxO activity and downregulated the expression of Pepck , thereby activating glycolysis and reducing glucose levels. By contrast, inhibiting insulin signaling increased the FoxO activity and upregulated the expression of Pepck , which activated gluconeogenesis and led to increased glucose levels. Our findings demonstrated the crucial role of the insulin signaling pathway in mediating glucose metabolism through the FoxO-Pepck axis, which supports the ecological adaptation of D. suzukii to high-sugar niches, thereby providing insights into its metabolic control and suggesting potential strategies for pest management. Elucidating these molecular processes is important for understanding metabolic regulation and ecological specialization in D. suzukii .

Both DNA 5-methylcytosine (5mC) and RNA N6-methyladenosine (m 6 A) modifications are reported to participate in cellular stress responses including inflammation. Phosphoenolpyruvate carboxykinase 2 (PCK2) is upregulated in Kupffer cells (KCs) to facilitate the proinflammatory phosphorylation signaling cascades upon LPS stimulation, yet the role of 5mC and m 6 A in PCK2 upregulation remain elusive. Here, we report that the significantly augmented PCK2 mRNA and protein levels are associated with global 5mC demethylation coupled with m 6 A hypermethylation in LPS-activated KCs. The suppression of 5mC demethylation or m 6 A hypermethylation significantly alleviates the upregulation of PCK2 and proinflammatory cytokines in LPS-challenged KCs. Further reciprocal tests indicate 5mC demethylation is upstream of m 6 A hypermethylation. Specifically, CpG islands in the promoters of PCK2 and RNA methyltransferase (METTL3 and METTL14) genes are demethylated, while the 3'UTR of PCK2 mRNA is m6A hypermethylated, in LPS-stimulated KCs. These modifications contribute to the transactivation of the PCK2 gene as well as increased PCK2 mRNA stability and protein production via a m 6 A-mediated mechanism with IGF2BP1 as the reader protein. These results indicate that DNA 5mC and RNA m6A collaborate to upregulate PCK2 expression, respectively, at the transcriptional and post-transcriptional levels during KC activation.

High dietary sugar (HDS), a contemporary dietary concern due to excessive intake of added sugars and carbohydrates, escalates the risk of metabolic disorders and concomitant cancers. However, the molecular mechanisms underlying HDS-induced cancer progression are not completely understood. We found that phosphoenolpyruvate carboxykinase 1 (PEPCK1), a pivotal enzyme in gluconeogenesis, is paradoxically upregulated in tumors by HDS, but not by normal dietary sugar (NDS), during tumor progression. Targeted knockdown of pepck1, but not pepck2, specifically in tumor tissue in Drosophila in vivo, not only attenuates HDS-induced tumor growth but also significantly improves the survival of Ras/Src tumor-bearing animals fed HDS. Interestingly, HP1a-mediated heterochromatin interacts directly with the pepck1 gene and downregulates pepck1 gene expression in wild-type Drosophila. Mechanistically, we demonstrated that, under HDS conditions, pepck1 knockdown reduces both wingless and TOR signaling, decreases evasion of apoptosis, reduces genome instability, and suppresses glucose uptake and trehalose levels in tumor cells in vivo. Moreover, rational pharmacological inhibition of PEPCK1, using hydrazinium sulfate, greatly improves the survival of tumor-bearing animals with pepck1 knockdown under HDS. This study is the first to show that elevated levels of dietary sugar induce aberrant upregulation of PEPCK1, which promotes tumor progression through altered cell signaling, evasion of apoptosis, genome instability, and reprogramming of carbohydrate metabolism. These findings contribute to our understanding of the complex relationship between diet and cancer at the molecular, cellular, and organismal levels and reveal PEPCK1 as a potential target for the prevention and treatment of cancers associated with metabolic disorders., (© 2024. The Author(s).)

Objectives: This study aimed to explore the involvement of phosphoenolpyruvate carboxykinase 2 (PCK2) in gefitinib-resistant non-small cell lung cancer (NSCLC) cells and assess its feasibility as a therapeutic target against gefitinib resistance., Methods: Gefitinib-resistant cell lines, PC9GR and HCC827GR, were generated through progressive exposure of parental cells to escalating concentrations of gefitinib. Transcriptomic analysis encompassed the treatment of PC9 and PC9GR cells with gefitinib or vehicle, followed by RNA extraction, sequencing, and subsequent bioinformatic analysis. Cell viability was determined via CCK-8 assay, while clonogenic assays assessed colony formation. Apoptosis was detected utilizing the Annexin V-FITC/7AAD kit. Iron ion concentrations were quantified using FerroOrange. mRNA analysis was conducted through quantitative RT-PCR. Western blotting was employed for protein analysis. H&E and immunohistochemical staining were performed on tumor tissue sections., Results: The results revealed that depletion or inhibition of PCK2 significantly enhanced gefitinib's efficacy in inducing cell growth arrest, apoptosis, and ferroptosis in resistant NSCLC. Moreover, PCK2 knockdown led to the downregulation of key ferroptosis-related proteins, GPX4 and SLC7A11, while upregulating ASCL4. Conversely, overexpression of PCK2 in gefitinib-sensitive cells rendered resistance to gefitinib. In vivo experiments using a gefitinib-resistant xenograft model demonstrated that PCK2 silencing not only reduced tumor growth but also considerably increased the anti-tumor effect of gefitinib., Conclusions: In conclusion, our study presents compelling evidence indicating that PCK2 plays a pivotal role in gefitinib resistance in NSCLC. The modulation of ferroptosis-related proteins and the involvement of Akt activation further elucidate the mechanisms underlying this resistance. Consequently, PCK2 emerges as a promising therapeutic target for overcoming gefitinib resistance in NSCLC, offering a new avenue for the development of more effective treatment strategies., 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.)

In adipose tissue, reduced expression of the glycerol channel aquaporin 7 (AQP7) has been associated with increased accumulation of triglyceride. The present study determines the relative protein abundances of lipolytic enzymes, AQP7, and cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) in paired mesenteric and omental visceral adipose tissue (VAT) and abdominal and femoral subcutaneous adipose tissue (SAT) in women with either normal weight or upper-body obesity. No differences in the expression of hormone-sensitive lipase (HSL) or AQP7 were found between the two groups in the four depots. The expression of adipocyte triglyceride lipase (ATGL) and HSL were higher in omental VAT and femoral SAT than in mesenteric VAT in both groups of women. Similarly, AQP7 expression was higher in omental VAT than in mesenteric VAT. The expression of PEPCK-C was lower in omental VAT than in femoral SAT. No correlation between the expression of AQP7 and the mean adipocyte size was observed; however, the expression of PEPCK-C positively correlated with the mean adipocyte size. In conclusion, a depot-specific protein expression pattern was found for ATGL, HSL, AQP7, and PEPCK-C. The expression pattern supports that the regulation of AQP7 protein expression is at least in part linked to the lipolytic rate. Furthermore, the results support that the synthesis of glycerol-3-phosphate via glyceroneogenesis contributes to regulating triglyceride accumulation in white adipose tissue in women.

Background: Phosphoenolpyruvate carboxykinase (PEPCK) plays a crucial role in gluconeogenesis, glycolysis, and the tricarboxylic acid cycle by converting oxaloacetate into phosphoenolpyruvate. Two distinct isoforms of PEPCK, specifically cytosolic PCK1 and mitochondrial PCK2, have been identified. Nevertheless, the comprehensive understanding of their dysregulation in pan-cancer and their potential mechanism contributing to signaling transduction pathways remains elusive., Methods: We conducted comprehensive analyses of PEPCK gene expression across 33 diverse cancer types using data from The Cancer Genome Atlas (TCGA). Multiple public databases such as HPA, TIMER 2.0, GEPIA2, cBioPortal, UALCAN, CancerSEA, and String were used to investigate protein levels, prognostic significance, clinical associations, genetic mutations, immune cell infiltration, single-cell sequencing, and functional enrichment analysis in patients with pan-cancer. PEPCK expression was analyzed about different clinical and genetic factors of patients using data from TCGA, GEO, and CGGA databases. Furthermore, the role of PCK2 in Glioma was examined using both in vitro and in vivo experiments., Results: The analysis we conducted revealed that the expression of PEPCK is involved in both clinical outcomes and immune cell infiltration. Initially, we verified the high expression of PCK2 in GBM cells and its role in metabolic reprogramming and proliferation in GBM., Conclusion: Our study showed a correlation between PEPCK (PCK1 and PCK2) expression with clinical prognosis, gene mutation, and immune infiltrates. These findings identified two possible predictive biomarkers across different cancer types, as well as a comprehensive analysis of PCK2 expression in various tumors, with a focus on GBM., Competing Interests: Declaration of competing interest The researchers declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Protein succinylation modification is a common post-translational modification (PTM) that plays an important role in bacterial metabolic regulation. In this study, quantitative analysis was conducted on the succinylated proteome of wild-type and florfenicol-resistant Vibrio alginolyticus to investigate the mechanism of succinylation regulating antibiotic resistance. Bioinformatic analysis showed that the differentially succinylated proteins were mainly enriched in energy metabolism, and it was found that the succinylation level of phosphoenolpyruvate carboxyl kinase (PEPCK) was highly expressed in the florfenicol-resistant strain. Site-directed mutagenesis was used to mutate the lysine (K) at the succinylation site of PEPCK to glutamic acid (E) and arginine (R), respectively, to investigate the function of lysine succinylation of PEPCK in the florfenicol resistance of V. alginolyticus . The detection of site-directed mutagenesis strain viability under florfenicol revealed that the survival rate of the E mutant was significantly higher than that of the R mutant and wild type, indicating that succinylation modification of PEPCK protein may affect the resistance of V. alginolyticus to florfenicol. This study indicates the important role of PEPCK during V. alginolyticus antibiotic-resistance evolution and provides a theoretical basis for the prevention and control of vibriosis and the development of new antibiotics.

Mitochondrial phosphoenolpyruvate carboxykinase (PCK2), a mitochondrial isoenzyme, supports the growth of cancer cells under glucose deficiency conditions in vitro. This study investigated the role and potential mechanism of PCK2 in the occurrence and development of Hepatocellular carcinoma (HCC). The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and other databases distinguish the expression of PCK2 and verified by qRT-PCR and Western blotting. Kaplan-Meier was conducted to assess PCK2 survival in HCC. The potential biological function of PCK2 was verified by enrichment analysis and gene set enrichment analysis (GSEA). The correlation between PCK2 expression and immune invasion and checkpoint was found by utilizing Tumor Immune Estimation Resource (TIMER). Lastly, the effects of PCK2 on the proliferation and metastasis of hepatocellular carcinoma cells were evaluated by cell tests, and the expressions of Epithelial mesenchymal transformation (EMT) and apoptosis related proteins were detected. PCK2 is down-regulated in HCC, indicating a poor prognosis. PCK2 gene mutation accounted for 1.3% of HCC. Functional enrichment analysis indicated the potential of PCK2 as a metabolism-related therapeutic target. Subsequently, we identified several signaling pathways related to the biological function of PCK2. The involvement of PCK2 in immune regulation was verified and key immune checkpoints were predicted. Ultimately, after PCK2 knockdown, cell proliferation and migration were significantly increased, and N-cadherin and vimentin expression were increased. PCK2 has been implicated in immune regulation, proliferation, and metastasis of hepatocellular carcinoma, and is emerging as a novel predictive biomarker and metabolic-related clinical target., (© 2024. The Author(s).)

In most actinomycetes, GlnR governs both nitrogen and non-nitrogen metabolisms (e.g., carbon, phosphate, and secondary metabolisms). Although GlnR has been recognized as a global regulator, its regulatory role in central carbon metabolism [e.g., glycolysis, gluconeogenesis, and the tricarboxylic acid (TCA) cycle] is largely unknown. In this study, we characterized GlnR as a direct transcriptional repressor of the pckA gene that encodes phosphoenolpyruvate carboxykinase, catalyzing the conversion of the TCA cycle intermediate oxaloacetate to phosphoenolpyruvate, a key step in gluconeogenesis. Through the transcriptomic and quantitative real-time PCR analyses, we first showed that the pckA transcription was upregulated in the glnR null mutant of Amycolatopsis mediterranei . Next, we proved that the pckA gene was essential for A. mediterranei gluconeogenesis when the TCA cycle intermediate was used as a sole carbon source. Furthermore, with the employment of the electrophoretic mobility shift assay and DNase I footprinting assay, we revealed that GlnR was able to specifically bind to the pckA promoter region from both A. mediterranei and two other representative actinomycetes ( Streptomyces coelicolor and Mycobacterium smegmatis ). Therefore, our data suggest that GlnR may repress pckA transcription in actinomycetes, which highlights the global regulatory role of GlnR in both nitrogen and central carbon metabolisms in response to environmental nutrient stresses., Importance: The GlnR regulator of actinomycetes controls nitrogen metabolism genes and many other genes involved in carbon, phosphate, and secondary metabolisms. Currently, the known GlnR-regulated genes in carbon metabolism are involved in the transport of carbon sources, the assimilation of short-chain fatty acid, and the 2-methylcitrate cycle, although little is known about the relationship between GlnR and the TCA cycle and gluconeogenesis. Here, based on the biochemical and genetic results, we identified GlnR as a direct transcriptional repressor of pckA , the gene that encodes phosphoenolpyruvate carboxykinase, a key enzyme for gluconeogenesis, thus highlighting that GlnR plays a central and complex role for dynamic orchestration of cellular carbon, nitrogen, and phosphate fluxes and bioactive secondary metabolites in actinomycetes to adapt to changing surroundings., Competing Interests: The authors declare no conflict of interest.

Backgruound: Sodium-dependent glucose cotransporter 2 (SGLT2) mediates glucose reabsorption in the renal proximal tubules, and SGLT2 inhibitors are used as therapeutic agents for treating type 2 diabetes mellitus. This study aimed to elucidate the effects and mechanisms of SGLT2 inhibition on hepatic glucose metabolism in both serum deprivation and serum supplementation states., Methods: Huh7 cells were treated with the SGLT2 inhibitors empagliflozin and dapagliflozin to examine the effect of SGLT2 on hepatic glucose uptake. To examine the modulation of glucose metabolism by SGLT2 inhibition under serum deprivation and serum supplementation conditions, HepG2 cells were transfected with SGLT2 small interfering RNA (siRNA), cultured in serum-free Dulbecco's modified Eagle's medium for 16 hours, and then cultured in media supplemented with or without 10% fetal bovine serum for 8 hours., Results: SGLT2 inhibitors dose-dependently decreased hepatic glucose uptake. Serum deprivation increased the expression levels of the gluconeogenesis genes peroxisome proliferator-activated receptor gamma co-activator 1 alpha (PGC-1α), glucose 6-phosphatase (G6pase), and phosphoenolpyruvate carboxykinase (PEPCK), and their expression levels during serum deprivation were further increased in cells transfected with SGLT2 siRNA. SGLT2 inhibition by siRNA during serum deprivation induces nuclear localization of the transcription factor forkhead box class O 1 (FOXO1), decreases nuclear phosphorylated-AKT (p-AKT), and p-FOXO1 protein expression, and increases phosphorylated-adenosine monophosphate-activated protein kinase (p-AMPK) protein expression. However, treatment with the AMPK inhibitor, compound C, reversed the reduction in the protein expression levels of nuclear p- AKT and p-FOXO1 and decreased the protein expression levels of p-AMPK and PEPCK in cells transfected with SGLT2 siRNA during serum deprivation., Conclusion: These data show that SGLT2 mediates glucose uptake in hepatocytes and that SGLT2 inhibition during serum deprivation increases gluconeogenesis via the AMPK/AKT/FOXO1 signaling pathway.