Mechanisms Of PUM2 Regulating Glioblastoma Cells Malignant Biological Behaviors

Objective: Glioblastoma is one of the most common central nervous system primary malignancy in adults.Though the biological behaviors of glioblastoma are characterized by rapid proliferation and aggressive local invasion in central nervous system,the molecular genetics of glioblastoma remains poorly understood,which impedes the development of more effective treatments.Revealing the molecular genetic mechanism of malignant biological behaviors in glioblastoma will contribute to developing new effective treatment options.PUM2,an RNA binding protein,is known to promote stem cell proliferation via repressing expressions of cell cycle genes.Similar with stem cells,malignant cells are characterized by unlimited proliferation and remote migration.However,roles of PUM2 in cancer development are controversial.BTG1 is a member of B cell translocation genes family.BTG1 has been proved to inhibite the activities of multiple non-central nervous system cancers,but the regulation of BTG1 in glioma is still unclear.Here,we investigated the role of PUM2 in the malignant biological behavior of glioma and its relationship with the cell cycle regulator BTG1,aiming to provide a new basis for the development of glioma and strive to provide new possible therapeutic targets.Methods: Immunoblotting and RT-qPCR were used to evaluate protein expression level and transcript level,respectively.ShRNAs were designed to knock down PUM2 and BTG1 expression.CCK-8 assay was used to evaluate cell viability.Transwell migration assay and evasion assay were used to evaluate metastatic capability of glioma cells.Annexin V/PI staining and flow cytometry assay were used to detect cell apoptosis.Actinomycin induction assay were used to measure the half-life of BTG1 mRNA.Double luciferase gene reporting system,RNA pull-down assay and RNA immunoprecipitation assay were used to test the interaction between PUM2 and BTG1 3′UTR.Results: PUM2 protein expression and gene transcription in glioma tissues was significantly higher than those in non-glioma tissues,and PUM2 protein expression in glioma cell lines was significantly higher than that in non-glioma cells.PUM2 expression was elevated in glioma tumor tissues as well as glioma cell lines.PUM2 knockdown remarkably suppressed glioma cell proliferation and migration,and also promoted glioma cell apoptosis.In addition,PUM2 knockdown increased BTG1 expression and prolonged half-life of BTG1 mRNA.PUM2 was binded to BTG1 3′UTR directly.Interestingly,BTG1 overexpression also suppressed glioma cell proliferation and migration,and promoted glioma cell apoptosis.Furthermore,BTG1 knockdown reversed the effect of PUM2 knockdown on glioma cell proliferation and migration.Conclusion: PUM2 expression is up-regulated in glioma tissues and cell lines.PUM2 is required for the proliferation,migration and invasion of glioma cells.Therefore,PUM2 are critical molecular characteristics of glioma cells.BTG1 is the critical target of PUM2 and its down regulation by PUM2 contributes to glioma development.PUM2 and BTG1 can be the potential pharmacotherapeutic targets in treatment of glioma.