Effects Of CDC73 On The Biological Behavior And Its Regulatory Mechanism In Glioblastoma

Objective: CDC73(cell division cycle 73 gene)has been widely studied as an oncogene,but an increasing number of studies have shown that CDC73 can also be converted into an oncogene under certain circumstances.Though its important roles in the development of tumors,there has not been any research reported in glioma.In this study,we used CDC73 in glioma tissues and glioblastoma(GBM)cell lines to initially verify the expression of CDC73 and analyze the correlation between its expression and the clinicopathological characteristics and prognosis of glioma patients based on clinical follow-up information;then we explored the effects of CDC73 on GBM proliferation,migration,apoptosis and cycle distribution through in vivo and in vitro experiments and its possible molecular mechanisms.Methods: Immunohistochemical staining was used to assess CDC73 expression in the glioma tissues and normal brain tissue.Combined with follow-up information,the relationship between the expression of CDC73 and clinicopathological features of patients,as well as the prognostic value were analyzed.The differential expression of CDC73 in GBM and its prognostic relevance were further analyzed by downloading TCGA data and online bioinformatic analysis tools.After detection of the expression of CDC73 by Western Blot and q RT-PCR,the GBM cell lines with high expression level of CDC73 were transfected with the constructed lentivirus vector containing sh CDC73 target sequence to establish GBM cell lines with low CDC73 expression.The effect of knockdown of CDC73 on the proliferation ability of GBM cell lines was examined by Cligo cell counting;changes in migration ability of GBM cell lines were assessed by scratch assay and transwell migration assay;effect on cycle distribution and apoptosis level of GBM cells before and after molecular intervention were assessed by flow cytometry.The molecules highly correlated with CDC73 were identified by Western Blot and verified by correlation analyse.Furthermore,the effects of downstream molecules were detected by reversion assay to explore the possible molecular mechanisms of CDC73 regulation of GBM development.Finally,rescue experiments were applied to confirm the results.The constructed CDC73 knockdown SHG-44 cell line was injected subcutaneously into BALB/c nude mice to establish a BALB/ C nude mouse xenograft tumor model to observe the effect of CDC73 on GBM growth in vivo.Results: 1.The expression of CDC73 in glioma tissue was significantly higher than that in normal brain tissue.Further SPSS survival analysis showed that the overall survival of glioma patients with high CDC73 expression was significantly shorter than that of those with low CDC73 expression.According to Mann-Whitney U analysis and Spearman rank correlation analysis,CDC73 expression was positively correlated with WHO grades and recurrence of glioma.Bioinformatic analysis also showed high expression of CDC73 in GBM.2.Knockdown of CDC73 significantly inhibited the proliferation and migration of GBM cells,promoted cell apoptosis and induce cell cycle rearrangement.3.The results of downstream molecular validation and reversion experiments showed that the expression of CDK1(cyclin-dependent kinases 1)was reduced after CDC73 knockdown;the effect of CDC73 on GBM cell migration and apoptosis could be rescued by down-regulating CDK1.4.In vivo experiments confirmed that knockdown of CDC73 inhibited tumor growth.Conclusion: CDC73 plays a vital role in the occurrence and development of gliomas.Additionally,CDC73 positively regulates the expression of CDK1,promotes the proliferation and migration of GBM cell lines,inhibits cell apoptosis,and promotes the cell cycle to stagnate in the G2/M phase.