Dataset Information

SNAP25 Inhibits Glioma Progression by Regulating Synapse Plasticity via GLS-Mediated Glutaminolysis.

ABSTRACT:

Background

Neuronal activity regulated by synaptic communication exerts an important role in tumorigenesis and progression in brain tumors. Genes for soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) annotated with the function 'vesicle' about synaptic connectivity were identified, and synaptosomal-associated protein 25 (SNAP25), one of those proteins, was found to have discrepant expression levels in neuropathies. However, the specific mechanism and prognostic value of SNAP25 during glioma progression remain unclear.

Methods

Using RNA sequencing data from The Cancer Genome Atlas (TCGA) database, the differential synaptosis-related genes between low grade glioma (LGG) and glioblastoma (GBM) were identified as highly correlated. Cox proportional hazards regression analysis and survival analysis were used to differentiate the outcome of low- and high-risk patients, and the Chinese Glioma Genome Atlas (CGGA) cohort was used for validation of the data set. RT-qPCR, western blot, and immunohistochemistry assays were performed to examine the expression level of SNAP25 in glioma cells and samples. Functional assays were performed to identify the effects of SNAP25 knockdown and overexpression on cell viability, migration, and invasion. Liquid chromatography-high resolution mass spectrometry (LC-MS)-based metabolomics approach was presented for identifying crucial metabolic disturbances in glioma cells. In situ mouse xenograft model was used to investigate the role of SNAP25 in vivo. Then, an immunofluorescence assay of the xenograft tissue was applied to evaluate the expression of the neuronal dendron formation marker-Microtubule Associated Protein 2 (MAP2).

Results

SNAP25 was decreased in level of expression in glioma tissues and cell lines, and low-level SNAP25 indicated an unfavorable prognosis of glioma patients. SNAP25 inhibited cell proliferation, migration, invasion and fostered glutamine metabolism of glioma cells, exerting a tumor suppressor role. Overexpressed SNAP25 exerted a lower expression level of MAP2, indicating poor neuronal plasticity and connectivity. SNAP25 could regulate glutaminase (GLS)-mediated glutaminolysis, and GLS knockdown could rescue the anti-tumor effect of SNAP25 in glioma cells. Moreover, upregulated SNAP25 also decreased tumor volume and prolonged the overall survival (OS) of the xenograft mouse.

Conclusion

SNAP25, a tumor suppressor inhibited carcinogenesis of glioma via limiting glutamate metabolism by regulating GLS expression, as well as inhibiting dendritic formation, which could be considered as a novel molecular therapeutic target for glioma.

SUBMITTER: Huang Q

PROVIDER: S-EPMC8416623 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:INTRODUCTION: Neuronal activity regulated by synaptic communication exerts an important role in tumorigenesis and progression in brain tumors. Genes for soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) annotated with the function ‘vesicle’ about synaptic connectivity were identified and one of these proteins, synaptosomal-associated protein 25 (SNAP25), was found to have discrepant expression levels in neuropathies. However, the specific mechanism and prognostic value of SNAP25 during glioma progression remain unclear. METHODS: Using RNA sequencing data from The Cancer Genome Atlas (TCGA) database, the differential synaptosis-related genes between LGG and GBM were identified as highly correlated. Cox proportional hazards regression analysis and survival analysis indicated that the candidate gene SNAP25 could differentiate the outcome of low- and high-risk patients, and the Chinese Glioma Genome Atlas (CGGA) cohort was used for validation of the data set. RT-qPCR, western blot, and immunohistochemistry assays were performed to examine the expression level of SNAP25 in glioma cells and samples. Functional assays were performed to identify the effects of SNAP25 knockdown and overexpression on cell viability, migration, and invasion. Then, an immunofluorescence assay of the xenograft tissue was applied to evaluate the expression of the neuronal dendron formation marker-MAP2. Liquid chromatography-high re solution mass spectrometry (LC-MS)-based metabolomics approach was presented for identifying crucial metabolic disturbances in glioma cells. In situ mouse xenograft model was used to investigate the role of SNAP25 in vivo.RESULTS: SNAP25 was down expressed in glioma tissues and cell lines and low-level SNAP25 indicated an unfavorable prognosis of glioma patients. SNAP25 inhibited cell proliferation, migration, invasion and fostered glutamate metabolism of glioma cells, exerting a tumor suppressor role. SNAP25 overexpression expressed lower expression of MAP2, indicating poor neuronal plasticity and connectivity. SNAP25 could interact with glutaminase（GLS）and GLS knockdown could rescue the anti-tumor effect of SNAP25 in glioma cells. Moreover, upregulation of SNAP25 also decreased tumor volume and prolonged the overall survival (OS) of the xenograft mouse.CONCLUSION: SNAP25 inhibited carcinogenesis of glioma via sponging glutamate metabolism by regulating GLS expression, as well as inhibiting dendritic formation, which could be considered as a molecular target for glioma diagnosis and therapy.

Project description:BackgroundA glioma is a tumor originating from glial cells in the central nervous system. Although significant progress has been made in diagnosis and treatment, most high-grade glioma patients are prone to recurrence. Therefore, molecular targeted therapy may become a new direction for adjuvant therapy in glioma. In recent years, many studies have revealed that circular RNA (circRNA) may play an important role in the occurrence and development of many tumors including gliomas. Our previous study found that the expression of hsa_circ_0008922 was up-regulated in glioma tissues upon RNA sequencing. The biological mechanism of circ_0008922 is still unreported in gliomas. Therefore, in this study, we preliminarily outlined the expression of hsa_circ_0008922 in glioma and explored its biological functions.MethodsThe expression of hsa_circ_0008922 in forty glioma tissues and four glioma cell lines (A172, U251, SF763 and U87) was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The correlation between hsa_circ_0008922 expression and clinicopathological features of glioma patients was evaluated by Fisher's exact test. To understand the potential function of hsa_circ_0008922 in glioma, we constructed small interfering RNA (siRNA) to hsa_circ_0008922 to downregulate its expression in glioma cell lines A172 and U251. With these hsa_circ_0008922 downregulated cells, a series of assays were carried out as follows. Cell proliferation was detected by CCK8 assay, migration and invasion were determined by wound healing assay and transwell assay, respectively. Colony formation ability was evaluated by plate clonogenic assay. Moreover, flow cytometry combined with Western blot was performed to analyze apoptosis status and the expression of apoptotic related proteins (caspase 3 and caspase 9). Finally, the possible biological pathways and potential miRNA targets of hsa_circ_0008922 were predicted by bioinformatics.ResultsWe found that the expression of hsa_circ_0008922 in glioma tissues was 3.4 times higher than that in normal tissues. The expression of has_circ_0008922 was correlated with WHO tumor grade. After down-regulating the expression of hsa_circ_0008922, malignant biological behavior of glioma cells was inhibited, such as cell proliferation, colony formation, migration, and invasion. At the same time, it also induced apoptosis of glioma cells. Predicted analysis by bioinformatics demonstrated that hsa_circ_0008922 may be involved in tumor-related pathways by acting as a molecular sponge for multiple miRNAs (hsa-let-7e-5p, hsa-miR-506-5p, hsa-let-7b-5p, hsa-let-7c-5p and hsa-let-7a-5p). Finally, we integrated our observation to build a circRNA-miRNA-mRNA predictive network.