Screening And Functional Verification Of Differentially Expressed Gene NUF2 In Breast Cancer

Objective:To screen differentially expressed gene in breast cancer,analyze its correlation with the prognosis of breast cancer,and study its effect on the proliferation and apoptosis of breast cancer cells.Methods:Bioinformatics analysis was performed using breast cancer-related microarray datasets in the GEO and TCGA databases to obtain differentially expressed genes(DEGs)in breast cancer;the biological functions and signaling pathways of DEGs were studied by GO and KEGG analyses.Through PPI network analysis and literature mining,NUF2 was selected as a research object;the expression of NUF2 in breast cancer tissues and its correlation with clinicopathological features and prognosis were analyzed using public database and clinical specimens;GSEA analysis predicts signaling pathways that NUF2 may be involved in.Two breast cancer cell lines,MCF-7 and MDA-MB-231 were transfected with NUF2 siRNA,and the transfection efficiency was verified by RT-qPCR and Western blot.The proliferation of breast cancer cells was detected by colony formation and CCK-8 assay.The apoptosis rate of breast cancer cells was detected by flow cytometry.Results:1.Using normal breast tissue as a control,190 DEGs with consistent expression trends were screened in GSE45827,GSE42568,GSE65194 and TCGA datasets,of which 65 were up-regulated DEGs and 125 were down-regulated DEGs.2.GO BP enrichment analysis suggested that up-regulated DEGs are mainly enriched in cell division,mitotic nuclear division,and G2/M transition of mitotic cell cycle,while downregulated DEGs were significantly associated with lipid metabolic process,cholesterol homeostasis,and glucose metabolic process.KEGG signaling pathway enrichment analysis indicated that up-regulated DEGs are associated with p53 signaling pathway,cell cycle and ECM-receptor interaction;while PPAR,AMPK signaling pathways are associated with down-regulated DEGs.3.Through PPI network analysis,a functional module containing 35 DEGs was chosen for further analysis.The 35 DEGs were ranked based on the log |FC| value of DEGs in the TCGA database,and literature mining of the top 10 DEGs.NUF2 was selected as a research ob.ject.4.NUF2 was up-regulated in breast cancer and was significantly associated with age(p<0.001),ER status(p<0.001),PR status(p<0.001),TNM stage(p=0.032)and molecular subtypes(p<0.001).5.NUF2 expression was significantly associated with breast cancer prognosis(p<0.05)and was more relevant in ER-positive breast cancer.With the increase of NUF2 expression,the overall survival and progression-free survival of breast cancer patients decreased.6.GSEA analysis showed that NUF2 is involved in the development of breast cancer mainly through cell cycle,DNA replication and p53 signaling pathway.7.After transfection of MCF-7 and MDA-MB-231 cells with NUF2 siRNA,the interference efficiency was confirmed by RT-qPCR and Western blot.Inhibition of the NUF2 expression can inhibit the proliferation ability and enhance apoptosis of the breast cancer cells.Conclusions:NUF2 was up-regulated in breast cancer and its expression level was significantly correlated with clinicopathological features such as ER status,PR status,TNM stage and molecular subtype.The prognosis of breast cancer patients with high NUF2 expression was poor,and NUF2 was expected to be a prognostic marker for breast cancer.Inhibition of the NUF2 expression can inhibit the proliferation ability and enhance apoptosis of the breast cancer cells,suggesting that NUF2 can be used as a potential therapeutic target for breast cancer.