MaHo001A: Identification of Novel Target Genes of Epstein-Barr Virus microRNAs

The human herpesvirus Epstein-Barr virus establishes persistent infections in the majority of the human population. Its main target cells are B lymphocytes and epithelial cells lining the oropharynx. Although primary infection usually occurs asymptomatically, EBV can cause mononucleosis. Moreover, EBV is strongly associated with multiple malignancies, including nasopharyngeal carcinoma, Burkitt Lymphoma, Hodgkin’s Lymphoma and post-transplant lymphoproliferative disease. In these EBV+ tumour tissues, the virus is latent and expresses only few proteins, yet up to ~40 miRNAs. Past research has indicated a possible causative link between these miRNAs and tumour development. However, the functions of these miRNAs remain poorly characterized. We aimed to explore the targetome of the EBV BART miRNAs by expressing these in a nasopharyngeal carcinoma cell line and performing whole genome gene expression profiling. Indeed, we identified numerous differentially expressed genes between miRNA-positive and -negative cells. By applying 3'UTR reporter assays, we validated eight genes as direct BART miRNA targets and identified the responsible miRNA(s). We found evidence for co-regulation of certain target genes by multiple miRNAs. Amongst these newly identified target genes was ATG16L1, a gene essential for autophagy. ATG16L1 protein levels were repressed by miR-BART18 in EBV+ Burkitt Lymphoma cells. HK-1 cells were lentivirally transduced to express the empty control vector (MH870) or a vector expressing the BART cluster 1 miRNAs (MH870-clus1). Sample versus common reference (cell line HK-1) hybridizations were performed on human whole genome gene expression microarrays V2 (Agilent, Belgium), in balanced dye-swap on 2 biological replicates per group.