A Preliminary Study Of Nasopharyngeal Carcinoma EBV-LMP1 By NF-κB Signaling Pathway In The Regulation Of WWOX

ObjectiveNasopharyngeal carcinoma (NPC) is the most frequent headand neck tumor in Guangdong, South China, and it is the mostcommonly known EBV associated cancer and expresses awell defined set of latent viral genes, including LMP1. Our previous studies found the expression of tumor suppressor gene WWOX was reduced in nasopharyngeal carcinoma, and WWOX expression gradually declined with the progress and lymph node metastasisin patients.These suggested thatWWOX was related with the clinical pathological featuresof nasopharyngeal carcinoma. LMP1 has been reported to be able to induce cell survival and growth throughNF-KB signaling pathway.This study mainly used oncogene LMP1 of EBV as a starting point to explore whether LMP1 regulated WWOXexpression through NF-κB signaling pathway.Methods1. Correlation between LMP1 and WWOX gene expression, NF-κB signaling pathwayWestern blot and qPCR were used to detect the expression of WWOX and NF-κB pathway (IκBα, p-IκBκ, p100/p52, p-p100/p52) in nasopharyngeal carcinoma cell lines CNE1 and CNE1-LMP1, and accessed relationship of LMP1 with NF-κB and WWOX.2. Studies of preliminary validation that LMP1 regulated WWOX expression by NF-κB pathwayWith proteasome inhibitors (MG132) of NF-kB signaling pathway to inhibit CNE1-LMP1 cell line, Western blot were used to detect the expression of WWOX and NF-κB pathway (IκBα, p-IκBκ, p100/p52, p-p100/p52) in the CNE1-LMP1 cell line, and verified that LMP1 regulated WWOX expression via NF-κB pathway.Results1. Correlation between LMP1 and WWOX gene, NF-κB signaling pathwayexpressionResearch of correlation betweenLMP1 and WWOXgene, protein expression:in CNE1-LMP1 cells, WWOX gene and protein levels were decreased compared with CNE1 cells (P=0.047, P=0.003, respectively), which was significantly different from each other, and suggested that LMP1 expression correlated with WWOX.Research of correlation between LMP1 and NF-κB signaling pathway: when nasopharyngeal carcinoma cell line CNE1-LMP1 was compared with CNE1 in NF-κB pathway key protein of IκBα, p-IκBα,p100/p52 and p-p100/p52 expression, P values were 0.06、0.012、0.083、0.038 respectively, and expression of p-IκBα, p-p100/p52 in CNE1-LMP1 were higher than CNE1, which were significantly different from each other. It suggested that NF-κB pathway was regulated by LMP1.2. Studies of preliminary validation that LMP1 regulated WWOX expression by NF-κB pathwayAfter using pathwayproteasome inhibitor (MG132) inhibits CNE1-LMP1 cell lines, expression levels of WWOX protein were elevated, P values were 0.043. It suggested that NF-κB signaling pathway regulated expression of WWOX gene. In addition, after using pathway proteasome inhibitor (MG132) inhibits, all P values of NF-κB signaling pathway key proteins (IκBα, p-IκB a, p100/p52, p-p100/p52) were 0.065、0.008、0.072、0.006, respectively. Protein expression of IκBα,p100/p52 was no statistically significant differencerespectively, while expression level of p-IκBα,p-p100/p52in CNE1-LMP1 cells was lower after drug inhibition. The difference was statistically significant. It indicated that the drug effectively inhibited protein expression of NF-κB pathway.Conclusions1. WWOX gene expression in CNE1-LMP1 cell lines was lower than CNE1 cell lines, which might be inhibited by the EBV-LMP1 oncogene.2.LMP1 could regulate NF-κB signaling pathway and its relevant expression of downstream genes.3. Reduced expression of WWOX might related with the regulation by LMP1 via NF-κB signaling pathway.