The ubiquitous Epstein-Barr virus (EBV) is associated with a subset of gastric carcinomas. MicroRNAs (miRs) are 22±2 nucleotide non-coding RNAs, and regulate various functions in cells. EBV-encoded miRs in gastric carcinoma cells have been identified, but the targets and roles of EBV-encoded miRs remain elusive. In the present study, we investigated cellular target of miR-BART5, and its possibility as an oncomiR. In naturally EBV-infected gastric carcinoma cells and EBV-positive gastric carcinoma tissues, the comprehensive EBV-miR profile revealed the expression of 22 EBV miRs (each having -3p and -5p) composed of miR-BART cluster 1 and cluster 2, without miR-BHRFs. Using bioinformatics analysis, we focused on miR-BART5 which shared seed sequence homology with hsa-miR-18a and miR-18b having oncomiR function, and protein inhibitor of activated STAT3 (PIAS3) mRNA. The western blot results confirmed that PIAS3 protein expression was reduced in miR-BART5-expressing gastric carcinoma cells (EBV-negative gastric carcinoma cells transfected with miR-BART5), compared with miR-control mock cells. Also PIAS3 expression was significantly lower in naturally EBV-infected gastric carcinoma cells than in EBV-negative gastric carcinoma or EBV-infected lymphoma cells. Moreover, there was a statistically significant inverse correlation between the expression of miR-BART5 and PIAS3. The luciferase reporter activity in cells diminished after co-transfection of pEZX-MT06 vector-PIAS3 3′UTR and miR-BART5, compared with co-transfection of empty vector and miR-control or co-transfection of empty vector and miR-BART5. When we tested twenty proteins as downstream candidates of miR-BART5, pSTAT3 increased and p21waf1 decreased in miR-BART5-expressing gastric carcinoma cells, compared with mock cells, and nuclear translocation of pSTAT3 was observed in miR-BART5-expressing gastric carcinoma cells. In a reverse context that naturally EBV-infected gastric carcinoma cells were transfected with anti-miR-BART5, protein levels were changed likewise; PIAS3 increased, pSTAT3 decreased and p21waf1 increased. However, there were no statistically significant differences in cellular biologic properties such as proliferation, apoptosis, invasion and migration between miR-BART5-expressing gastric carcinoma cells and mock cells, or between naturally EBV-infected gastric carcinoma cells transfected with anti-miR-BART5 and mock cells. Taken together, miR-BART5 not only targets directly PIAS3 and then modulates PIAS3-pSTAT3 axis, but also decreases p21waf1 protein, although it fails to alter cellular proliferation in gastric carcinoma cells. This suggests that a single viral microRNA is not sufficient to implement oncogenic function, despite directly targeting cellular protein involved in oncogenesis. Instead, a whole cluster like miR-BART cluster 1 or cluster 2 may be advocate to control cellular biologic property.

Citation Format: Dong Ha Kim, Chan Jin Yoon, Jin-Seoub Kim, Sunyoung Park, Euno Choi, Jun Hee Woo, Mee Soo Chang. Epstein-Barr virus-encoded miR-BART5 modulates PIAS3-pSTAT3 and p21waf1 in gastric carcinoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 470. doi:10.1158/1538-7445.AM2017-470