SNAP25 Inhibits Glioma Progression Via Glutamate Mediated Neuro-glioma Synaptogenesis

BackgroundNeuronal activity regulated by synaptic communication exerts an important role in cell proliferation,migration and invasion in brain tumors.Genes for soluble Nethylmaleimide-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.ObjectiveFirstly,we evaluated expression of SNAP25 in glioma cell lines,glioma tissues and estimated its clinical relevance.Next,we explored the role of SNAP25 and its target gene GLS on glioma cell proliferation,cell migration and invasion.Then,we studied the metabolical and neuronal formation function of SNAP25.Finally,we test the hypothesis that SNAP25 degradation enhanced glioma progression and glutamine metabolism by sponging glutaminase（GLS）expression in glioma cells.MethodsEleven synapse formation-related genes were profiled using RNA sequencing data from The Cancer Genome Atlas（TCGA）database.According to the differential genes between LGG and GBM,we developed an eleven-gene set using Cox proportional hazards regression analysis,and Chinese Glioma Genome Atlas（CGGA）cohort was used for validation of the data set.Quantitative RT-PCR,western-blot and immunohistochemistry assays were performed to examine the expression level of SNAP25 in glioma cells and samples.The CCK-8,wound healing and transwell assays were performed to identify the effects of SNAP25 knockdown and overexpression on cell viability,apoptosis,migration and invasion,respectively.Then,immunofluorescence assay of the xenograft tissue was applied to evaluate the expression of the neuronal dendron formation marker—MAP2.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 xenograft glioma growth.ResultsSNAP25 was down-expressed in glioma tissues and cell lines,and low-level of SNAP25 indicated unfavorable prognosis of glioma patients.Overexpressed SNAP25 inhibited cell proliferation,migration,invasion and fostered glutamine metabolism of glioma cells,exerting a tumor suppressor role.SNAP25 overexpression expressed lower MAP2,indicating poor neuronal plasticity and connectivity.SNAP25 could interact with glutaminase（GLS）and GLS knockdown could rescue the anti-tumor effect of SNAP25 on glioma cells.Moreover,upregulation of SNAP25 also decreased tumor volume and prolonged the overall survival of the xenograft mouse in vivo.ConclusionSNAP25 inhibited carcinogenesis progression of glioma via targeting glutamine metabolism by regulating GLS expression,as well as inhibiting dendron formation,which could be considered as a molecular target for glioma diagnosis and therapy.