Preliminary Study Of Differential Gene Expression Analysis Of EB Virus-associated Burkitt’s Lymphoma

Objective: At the molecular level,the Epstein-Barr Virus(EBV)-related differentially expressed genes(DEGs)of Burkitt Lymphoma(BL)were explored to provide new ideas for its diagnosis and treatment.To clarify the expression of pre-m RNA processing factor 19(PRPF19),UBE2 N,CDH1in Epstein-Barr virus-associated Burkitt’s lymphoma cell lines and tissues,to explore the causes of differences in differentially expressed gene expression and the transcription by EBV-encoded latent genes.To explore the interaction between EBV and differentially expressed gene in the development of EBV-associated Burkitt’s lymphoma,and to provide experimental evidence for elucidating the role of differentially expressed gene in Burkitt’s lymphoma.Methods: Data set GSE100458 for EBV-positive and negative BL-related gene chips was obtained from the National Center for Biotechnology Information(NCBI).Using Morpheus software screens for differentially expressed genes,enter the gene for up-and down-regulation into DAVID database separately and analysis them using GO.Further,10 TOP genes were screened out using cytoscape software.Quantitative Real-time PCR(q RT-PCR)was used to detect the transcriptional expression of PRPF19 and other genes in EBV-positive lymphoma cell lines Raji,Daudi and EBV-negative lymphoma cell line Ramos.Bisulfite genomic sequencing(BGS)was used to detect the methylation status of the promoter region of PRPF19 gene in EBV-positive Burkitt’s lymphoma cell line and EBV-negative Burkitt’s lymphoma cell line.The expression of PRPF19 and other genes in EBV-positive Burkitt’s lymphoma cell line and EBV-negative Burkitt’s lymphoma cell line were detected by immunohistochemistry.Statistical analysis was performed using t test and Fisher’s exact test.Results: The first 400 differentially expressed genes in the two groups were analyzed,including 200 up-regulated genes and 200 down-regulated genes.The up-regulated genes were mainly concentrated in the extracellular space pathway,heparin-binding pathway,and growth factor activity pathway.Down-regulated genes are mainly concentrated in retrograde small vesicle trafficking pathway,endoplasmic reticulum pathway,and gamma tubulin binding pathway.After the differential gene was analyzed by protein interaction,10 genes including the core gene PRPF19 were screened out.The results of real-time PCR showed that there were statistical differences in the transcriptional expression of 7 genes.There is no significant difference in the positive rate of PRPF19 protein between Burkitt’s lymphoma-positive tissue and Burkitt’s lymphoma-negative tissue,suggesting that there may be other more complicated regulatory pathways for PRPF19 expression in Burkitt’s lymphoma.Continue to increase the sample size for further study.including PRPF19 between EBV positive and negative BL groups.There were no statistics on three genes including ACACB(acetyl-Co A carboxylase beta).The methylation status of the promoter regions of the gene PRPF19 and UBE2 N was detected.The BGS sequencing results showed that neither the EBV-positive BL cell line nor the EBV-negative BL cell line had methylation in the PRPF19 and UBE2 N promoter regions.UBE2 N,CDH1,and PRPF19 genes were selected for protein level verification.Western blotting showed differential expression of UBE2 N in EBV-positive and negative BL lymphoma cell lines(t=18.56,P=0.0029).The expression level of PRPF19 was higher,and there was no statistical difference(t=0.5369,P=0.6286),and CDH1 was not expressed at all.Conclusion:(1)The transcription level of PRPF19 and UBE2 N in EBV-positive BL cell line was significantly lower than that in EBV-negative BL cell line.The transcription level of CDH1 in EBV-positive BL cell line was significantly higher than that in EBV-negative BL cell line,suggesting that EBV infection can regulate the transcription level of differential gene.(2)The methylation status of PRPF19 and UBE2 N promoter regions in EBV-positive BL and EBV-negative BL cell lines did not change,suggesting that the down-regulation of differential gene transcription levels in EBV-positive BL is not caused by promoter methylation.(3)There was a significant difference in the positive rate of UBE2 N protein between BL-positive and BL-negative tissues.There was no significant difference in the positive rate of PRPF19 protein between BL-positive and BL-negative tissues,which may be related to the BL samples in this study,perhaps in BL tissues.There may be other more complicated regulatory pathways for differential gene expression.(4)EBV infection can lead to changes in the gene expression profile of host cells.Bioinformatics analysis can initially screen differentially expressed genes and interacting proteins,providing valuable information and ideas for further research.