Epstein-Barr virus-coded miR-BART13 promotes nasopharyngeal carcinoma cell growth and metastasis via targeting of the NKIRAS2/NF-κB pathway.

Based on analysis of Epstein-Barr virus (EBV) BART microRNA expression profiles, we previously reported that EBV-encoded miR-BART13 is upregulated in nasopharyngeal carcinoma (NPC) plasma specimens. However, the effects and molecular mechanisms of miR-BART13 in NPC remain largely unknown. We found that miR-BART13 was significantly upregulated in NPC tissue specimens. Ectopic expression of miR-BART13 promoted NPC cell proliferation, epithelial mesenchymal transition, and metastasis in vitro, and facilitated xenograft tumor growth and lung metastasis in vivo. Molecularly, NF-κB inhibitor interacting Ras-like 2 (NKIRAS2), a negative regulator of the NF-κB signaling, was identified to be a direct target of miR-BART13 in NPC cells, and NKIRAS2 mRNA and protein expression was inversely correlated with miR-BART13 in NPC tissues, respecitvely. Furthermore, the NF-κB signaling pathway was activated by miR-BART13. By rescued experiments, reconstitution of NKIRAS2 expression abrogated all the phenotypes upregulated by miR-BART13, and attenuated activity of NF-κB signaling pathway activated by miR-BART13 in NPC cells. Our findings indicated the newly identified miR-BART13/NKIRAS2/NF-κB signaling axis may provide further insights into better understanding of NPC initiation and development, and targeting of this pathway could be further studied as a therapeutic strategy for NPC patients.

Xu YJ ，Zhou R ，Zong JF ，Lin WS ，Tong S ，Guo QJ ，Lin C ，Lin SJ ，Chen YX ，Chen MR ，Chen HL ，Ye YB ，Pan JJ ... -  《-》

EBV-miR-BART8-3p induces epithelial-mesenchymal transition and promotes metastasis of nasopharyngeal carcinoma cells through activating NF-κB and Erk1/2 pathways.

Lin C ，Zong J ，Lin W ，Wang M ，Xu Y ，Zhou R ，Lin S ，Guo Q ，Chen H ，Ye Y ，Zhang B ，Pan J ... -  《JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH》

Circulating Epstein-Barr virus microRNAs miR-BART7 and miR-BART13 as biomarkers for nasopharyngeal carcinoma diagnosis and treatment.

More than 75% of nasopharyngeal carcinoma (NPC) patients have already developed local or regional spread at diagnosis, which hampers effective treatment and results in a poor prognosis. It is essential to characterize more sensitive and specific biomarkers for screening of high risk individuals and assessment of NPC treatment effectiveness. NPC is an Epstein-Barr virus (EBV) associated tumor in which only a few viral proteins but more than 20 BamHI A rightward transcripts (BART) microRNAs are detected, at abundant levels. We hypothesized that these BART microRNAs may be novel biomarkers for NPC. Systematic analysis of EBV BART microRNA expression profiles in EBV latently infected Mutu I and Mutu III cell lines, EBV-harboring NPC and noncancerous NP cells found that miR-BART3, miR-BART7 and miR-BART13 microRNAs are highly expressed and regularly secreted into the extracellular environment of NPC cells. These BART microRNAs were evaluated for used as potential NPC biomarkers. Analysis of plasma specimens obtained from NPC patients (n = 89), and healthy (n = 28) and non-NPC tumor patient controls (n = 18) found levels of both miR-BART7 and miR-BART13, but not miR-BART3, to be distinctly presence among NPC patients, with elevated levels being particularly apparent among patients with advanced disease. Receiver operating characteristic curve analysis combining miR-BART7 and miR-BART13 levels produces a 90% predictive value for the presence of NPC. Analysis of 41 NPC patients before and after radiotherapy showed that miR-BART7 and miR-BART13, but not miR-BART3, were diminished after treatment. These results indicate that EBV microRNAs, miR-BART7 and miR-BART13, may constitute useful new serological biomarkers for diagnosis of NPC and prediction of treatment efficacy.

Zhang G ，Zong J ，Lin S ，Verhoeven RJ ，Tong S ，Chen Y ，Ji M ，Cheng W ，Tsao SW ，Lung M ，Pan J ，Chen H ... -  《-》

NF-κB Signaling Regulates Expression of Epstein-Barr Virus BART MicroRNAs and Long Noncoding RNAs in Nasopharyngeal Carcinoma.

Epstein-Barr virus (EBV) expresses few viral proteins in nasopharyngeal carcinoma (NPC) but high levels of BamHI-A rightward transcripts (BARTs), which include long noncoding RNAs (lncRNAs) and BART microRNAs (miRNAs). It is hypothesized that the mechanism for regulation of BARTs may relate to EBV pathogenesis in NPC. We showed that nuclear factor-κB (NF-κB) activates the BART promoters and modulates the expression of BARTs in EBV-infected NPC cells but that introduction of mutations into the putative NF-κB binding sites abolished activation of BART promoters by NF-κB. Binding of p50 subunits to NF-κB sites in the BART promoters was confirmed in electrophoretic mobility shift assays (EMSA) and further demonstrated in vivo using chromatin immunoprecipitation (ChIP) analysis. Expression of BART miRNAs and lncRNAs correlated with NF-κB activity in EBV-infected epithelial cells, while treatment of EBV-harboring NPC C666-1 cells with aspirin (acetylsalicylic acid [ASA]) and the IκB kinase inhibitor PS-1145 inhibited NF-κB activity, resulting in downregulation of BART expression. Expression of EBV LMP1 activates BART promoters, whereas an LMP1 mutant which cannot induce NF-κB activation does not activate BART promoters, further supporting the idea that expression of BARTs is regulated by NF-κB signaling. Expression of LMP1 is tightly regulated in NPC cells, and this study confirmed that miR-BART5-5p downregulates LMP1 expression, suggesting a feedback loop between BART miRNA and LMP1-mediated NF-κB activation in the NPC setting. These findings provide new insights into the mechanism underlying the deregulation of BARTs in NPC and identify a regulatory loop through which BARTs support EBV latency in NPC. Nasopharyngeal carcinoma (NPC) cells are ubiquitously infected with Epstein-Barr virus (EBV). Notably, EBV expresses very few viral proteins in NPC cells, presumably to avoid triggering an immune response, but high levels of EBV BART miRNAs and lncRNAs which exhibit complex functions associated with EBV pathogenesis. The mechanism for regulation of BARTs is critical for understanding NPC oncogenesis. This study provides multiple lines of evidence to show that expression of BARTs is subject to regulation by NF-κB signaling. EBV LMP1 is a potent activator of NF-κB signaling, and we demonstrate that LMP1 can upregulate expression of BARTs through NF-κB signaling and that BART miRNAs are also able to downregulate LMP1 expression. It appears that aberrant NF-κB signaling and expression of BARTs form an autoregulatory loop for maintaining EBV latency in NPC cells. Further exploration of how targeting NF-κB signaling interrupts EBV latency in NPC cells may reveal new options for NPC treatment.

Verhoeven RJ ，Tong S ，Zhang G ，Zong J ，Chen Y ，Jin DY ，Chen MR ，Pan J ，Chen H ... -  《-》

Downregulation of ABI2 expression by EBV-miR-BART13-3p induces epithelial-mesenchymal transition of nasopharyngeal carcinoma cells through upregulation of c-JUN/SLUG signaling.

Huang J ，Qin Y ，Yang C ，Wan C ，Dai X ，Sun Y ，Meng J ，Lu Y ，Li Y ，Zhang Z ，Wu B ，Xu S ，Jin H ，Yang K ... -  《Aging-US》