Analysis of the expression, function and signaling of glycogen phosphorylase isoforms in hepatocellular carcinoma.
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SUBMITTER: Jiang L
PROVIDER: S-EPMC9214699 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background and purposeGlycogen phosphorylase (GP) is the key enzyme for glycogen degradation. GP inhibitors (GPi-s) are glucose lowering agents that cause the accumulation of glucose in the liver as glycogen. Glycogen metabolism has implications in beta cell function. Glycogen degradation can maintain cellular glucose levels, which feeds into catabolism to maintain insulin secretion, and elevated glycogen degradation levels contribute to glucotoxicity. The purpose of this study was to assess whether influencing glycogen metabolism in beta cells by GPi-s affects the function of these cells.Experimental approachThe effects of structurally different GPi-s were investigated on MIN6 insulinoma cells and in a mouse model of diabetes.Key resultsGPi treatment increased glycogen content and, consequently, the surface area of glycogen in MIN6 cells. Furthermore, GPi treatment induced insulin receptor β (InsRβ), Akt and p70S6K phosphorylation, as well as pancreatic and duodenal homeobox 1(PDX1) and insulin expression. In line with these findings, GPi-s enhanced non-stimulated and glucose-stimulated insulin secretion in MIN6 cells. The InsRβ was shown to co-localize with glycogen particles as confirmed by in silico screening, where components of InsR signalling were identified as glycogen-bound proteins. GPi-s also activated the pathway of insulin secretion, indicated by enhanced glycolysis, mitochondrial oxidation and calcium signalling. Finally, GPi-s increased the size of islets of Langerhans and improved glucose-induced insulin release in mice.Conclusion and implicationsThese data suggest that GPi-s also target beta cells and can be repurposed as agents to preserve beta cell function or even ameliorate beta cell dysfunction in different forms of diabetes.Linked articlesThis article is part of a themed section on Inventing New Therapies Without Reinventing the Wheel: The Power of Drug Repurposing. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.2/issuetoc.
| S-EPMC5758390 | biostudies-literature
Project description:BACKGROUND: Mutations in the PYGM gene encoding skeletal muscle glycogen phosphorylase (GP) cause a metabolic disorder known as McArdle's disease. Previous studies in muscle biopsies and cultured muscle cells from McArdle patients have shown that PYGM mutations abolish GP activity in skeletal muscle, but that the enzyme activity reappears when muscle cells are in culture. The identification of the GP isoenzyme that accounts for this activity remains controversial. METHODOLOGY/PRINCIPAL FINDINGS: In this study we present two related patients harbouring a novel PYGM mutation, p.R771PfsX33. In the patients' skeletal muscle biopsies, PYGM mRNA levels were ?60% lower than those observed in two matched healthy controls; biochemical analysis of a patient muscle biopsy resulted in undetectable GP protein and GP activity. A strong reduction of the PYGM mRNA was observed in cultured muscle cells from patients and controls, as compared to the levels observed in muscle tissue. In cultured cells, PYGM mRNA levels were negligible regardless of the differentiation stage. After a 12 day period of differentiation similar expression of the brain and liver isoforms were observed at the mRNA level in cells from patients and controls. Total GP activity (measured with AMP) was not different either; however, the active GP activity and immunoreactive GP protein levels were lower in patients' cell cultures. GP immunoreactivity was mainly due to brain and liver GP but muscle GP seemed to be responsible for the differences. CONCLUSIONS/SIGNIFICANCE: These results indicate that in both patients' and controls' cell cultures, unlike in skeletal muscle tissue, most of the protein and GP activities result from the expression of brain GP and liver GP genes, although there is still some activity resulting from the expression of the muscle GP gene. More research is necessary to clarify the differential mechanisms of metabolic adaptations that McArdle cultures undergo in vitro.
| S-EPMC2950139 | biostudies-literature
Project description:Inhibition of glycogen breakdown blocks memory formation in young animals, but it stimulates the maintenance of the long-term potentiation, a cellular mechanism of memory formation, in hippocampal slices of old animals. Here, we report that a 2-week treatment with glycogen phosphorylase inhibitor BAY U6751 alleviated memory deficits and stimulated neuroplasticity in old mice. Using the 2-Novel Object Recognition and Novel Object Location tests, we discovered that the prolonged intraperitoneal administration of BAY U6751 improved memory formation in old mice. This was accompanied by changes in morphology of dendritic spines in hippocampal neurons, and by "rejuvenation" of hippocampal proteome. In contrast, in young animals, inhibition of glycogen degradation impaired memory formation; however, as in old mice, it did not alter significantly the morphology and density of cortical dendritic spines. Our findings provide evidence that prolonged inhibition of glycogen phosphorolysis improves memory formation of old animals. This could lead to the development of new strategies for treatment of age-related memory deficits.
| S-EPMC10497847 | biostudies-literature
Project description:BackgroundExcessive hepatic glycogen accumulation benefits tumorigenesis and cancer cell survival. We previously reported that BMP4 has the strongest ability to promote glycogenesis among the 14 BMPs in hepatocytes and augmented hepatocellular carcinoma (HCC) cell survival under hypoxia and hypoglycemia conditions by promoting the glycolysis pathway. However, the mechanism underlying BMP4's effect on glycogenesis in HCC remains elusive.MethodsThe expression of BMP4 and SLC2A1 were acquired by analyzing the TCGA-LIHC dataset, as well as by immunohistochemical analysis of the 40 pairs of human HCC samples and para-tumor tissues. Gene expressions were detected by qPCR, immunoflurorescence staining, and Western blotting. Overexpression and silencing of BMP4 were accomplished through adenoviruses Ad-B4 and Ad-siB4 infection. Hepatic glycogen was detected by PAS staining. SLC2A1 (GLUT1) function was blocked by the inhibitor BAY-876. ChIP assay was used to determine the binding of SMADs to the promoter region of SLC2A1 in HCC cells. Lastly, the in vivo effect of BMP4-regulated SLC2A1 on HCC tumor growth was assessed in a xenograft model of HCC.ResultsThe elevated expression of BMP4 in HCC tumor tissues was highly correlated with hepatic glycogen accumulation in clinical samples. SLC2A1 was highly expressed in HCC tumor tissue and correlated with clinical stage and prognosis. Exogenous BMP4 augmented glycogen accumulation and upregulated the expression of glycogen synthesis-related genes in Huh7 and HepG2 cells, both of which were effectively blunted by SLC2A1inhibitor BAY-876. In mechanism, BMP4 activated SMAD5 to regulate the promoter of SLC2A1to enhance its expression. The in vivo xenograft experiments revealed that BMP4 promoted glycogen accumulation and tumor growth, which were effectively diminished by BAY-876.ConclusionThese results demonstrate that BMP4 upregulates glycogen synthesis through the SMAD/SLC2A1 (GLUT1) signaling axis in HCC cells, which may be exploited as novel therapeutic targets for HCC treatment.
| S-EPMC10339511 | biostudies-literature
Project description:Brain glycogen metabolism plays a critical role in major brain functions such as learning or memory consolidation. However, alteration of glycogen metabolism and glycogen accumulation in the brain contributes to neurodegeneration as observed in Lafora disease. Glycogen phosphorylase (GP), a key enzyme in glycogen metabolism, catalyzes the rate-limiting step of glycogen mobilization. Moreover, the allosteric regulation of the three GP isozymes (muscle, liver, and brain) by metabolites and phosphorylation, in response to hormonal signaling, fine-tunes glycogenolysis to fulfill energetic and metabolic requirements. Whereas the structures of muscle and liver GPs have been known for decades, the structure of brain GP (bGP) has remained elusive despite its critical role in brain glycogen metabolism. Here, we report the crystal structure of human bGP in complex with PEG 400 (2.5 Å) and in complex with its allosteric activator AMP (3.4 Å). These structures demonstrate that bGP has a closer structural relationship with muscle GP, which is also activated by AMP, contrary to liver GP, which is not. Importantly, despite the structural similarities between human bGP and the two other mammalian isozymes, the bGP structures reveal molecular features unique to the brain isozyme that provide a deeper understanding of the differences in the activation properties of these allosteric enzymes by the allosteric effector AMP. Overall, our study further supports that the distinct structural and regulatory properties of GP isozymes contribute to the different functions of muscle, liver, and brain glycogen.
| S-EPMC5000057 | biostudies-literature
Project description:RNA interference has been used to study insects' gene function and regulation. Glycogen synthase (GS) and glycogen phosphorylase (GP) are two key enzymes in carbohydrates' conversion in insects. Glycogen content and GP and GS gene expression in several tissues and developmental stages of the Brown planthopper Nilaparvata lugens Stål (Hemiptera: Delphacidae) were analyzed in the present study, using quantitative reverse-transcription polymerase chain reaction to determine their response to double-stranded trehalases (dsTREs), trehalose-6-phosphate synthases (dsTPSs), and validamycin injection. The highest expression of both genes was detected in the wing bud, followed by leg and head tissues, and different expression patterns were shown across the developmental stages analyzed. Glycogen content significantly decreased 48 and 72?h after dsTPSs injection and 48?h after dsTREs injection. GP expression increased 48?h after dsTREs and dsTPSs injection and significantly decreased 72?h after dsTPSs, dsTRE1-1, and dsTRE1-2 injection. GS expression significantly decreased 48?h after dsTPS2 and dsTRE2 injection and 72?h after dsTRE1-1 and dsTRE1-2 injection. GP and GS expression and glycogen content significantly decreased 48?h after validamycin injection. The GP activity significantly decreased 48?h after validamycin injection, while GS activities of dsTPS1 and dsTRE2 injection groups were significantly higher than that of double-stranded GFP (dsGFP) 48?h after injection, respectively. Thus, glycogen is synthesized, released, and degraded across several insect tissues according to the need to maintain stable trehalose levels.
| S-EPMC5469382 | biostudies-literature
Project description:The selective phosphorylation of glycogen phosphorylase (GP) by its only known kinase, phosphorylase kinase (PhK), keeps glycogen catabolism tightly regulated. In addition to the obligatory interaction between the catalytic ? subunit of PhK and the phosphorylatable region of GP, previous studies have suggested additional sites of interaction between this kinase and its protein substrate. Using short chemical crosslinkers, we have identified direct interactions of GP with the large regulatory ? and ? subunits of PhK. These newfound interactions were found to be sensitive to ligands that bind PhK.
| S-EPMC5195864 | biostudies-literature
Project description:Tumor progression is dependent on metabolic reprogramming. Metastasis and vasculogenic mimicry (VM) are typical characteristics of tumor progression. The relationship among metastasis, VM, and metabolic reprogramming remains unclear. In this study, we identified the novel role of Twist1, a VM regulator, in the transcriptional regulation of thymidine phosphorylase (TP) expression. TP promoted the extracellular metabolism of thymidine into ATP and amino acids through the pentose Warburg effect by coupling the pentose phosphate pathway and glycolysis. Moreover, Twist1 relied on TP-induced metabolic reprogramming to promote hepatocellular carcinoma (HCC) metastasis and VM formation mediated by VE-Cad, VEGFR1, and VEGFR2 in vitro and in vivo. The TP inhibitor tipiracil reduced the effect of TP on promoting HCC VM formation and metastasis. Hence, TP, when transcriptionally activated by Twist1, promotes HCC VM formation and metastasis through the pentose Warburg effect and contributes to tumor progression.
| S-EPMC6426839 | biostudies-literature
Project description:The spleen tyrosine kinase (SYK) has been reported as a novel biomarker for human hepatocellular carcinoma, but the functional contributions of its two isoforms SYK(L) and SYK(S) are undefined. In this study, we investigated their biologic functions and possible prognostic values in hepatocellular carcinoma. SYK(L) was downregulated in 38% of human specimens of hepatocellular carcinoma examined, whereas SYK(S) was detectable in 40% of these specimens but not in normal liver tissue samples without cirrhosis. SYK(S) expression correlated with pathologic parameters characteristic of tumor metastasis, including multiple tumors (P = 0.003) and vascular invasion (P = 0.001). Further, SYK(S) was specifically associated with epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma specimens. Functional studies showed that SYK(S) promoted tumor growth, suppressed apoptosis, and induced EMT through the extracellular signal-regulated kinase pathway, countering the opposite effects of SYK(L). Patients with SYK(L(+)/S(-)) tumors exhibited longer overall survival and time to recurrence than those with SYK(L(-)/S(-)) or SYK(L(+)/S(+)) tumors (P < 0.001). Taken together, our findings showed that SYK(S) enhances invasion, whereas SYK(L) inhibits metastasis in hepatocellular carcinoma. We suggest that SYK(L) downregulation or SYK(S) elevation are strong predictors of poor survival in patients with hepatocellular carcinoma, indicative of a need for aggressive therapeutic intervention.
| S-EPMC3959256 | biostudies-literature
Project description:BACKGROUND: Primary hepatocellular carcinoma (HCC) is one of the most common malignancies in the world. However, the molecular pathogenesis of HCC is not well-understood, and the prognosis for patients with HCC remains very poor. METHODS: To disclose detailed genetic mechanisms in hepatocellular carcinoma (HCC) with a view toward development of novel therapeutic targets, we analyzed expression profiles HCCs and their corresponding noncancerous tissues by using bioinformatics method. RESULTS: In this paper, we report the identification of genes whose expression has been altered and the changed bio-pathways during hepatocarcinogenesis. Hepatoma cells infect intracellular and intercellular signal transduction through Focal adhesion and cause abnormal expression of important intracellular signaling pathway. In addition, it is worth mentioning that some small molecules still restored to the state similar to normal cells, such as bambuterol and lovastatin. This member gene set would serve as a pool of lead gene targets for the identification and development of novel diagnostic and therapeutic biomarkers to greatly improve the clinical management of HCC patients with different risks of recurrence after curative partial hepatectomy. CONCLUSIONS: The study has great significance for gene therapy and pharmacotherapy and provides a new treatment entry point and a potential new clinical drug for HCC patients.
| S-EPMC4177135 | biostudies-literature