Identification of new viral genes and transcript isoforms during Epstein-Barr virus reactivation using RNA-seq.

Using an enhanced RNA-seq pipeline to analyze Epstein-Barr virus transcriptomes, we have investigated viral and cellular gene expression in the Akata cell line following B-cell receptor mediated reactivation. Robust induction of EBV gene expression was observed, with most viral genes induced more than 200-fold and with EBV transcripts accounting for 7% of all mapped reads within the cell. Following induction, hundreds of candidate splicing events were detected using the junction mapper TopHat, including a novel non-productive splicing event at the gp350/gp220 locus and several alternative splicing events at the LMP2 locus. A more detailed analysis of lytic LMP2 transcripts showed an overall lack of the prototypical type III latency splicing events. Analysis of nuclear versus cytoplasmic RNA-seq data showed that the lytic forms of LMP2, EBNA-2, EBNA-LP, and EBNA-3A, B, and C have higher nuclear-to-cytoplasmic accumulation ratios than most lytic genes, including classic late genes. This data raises the possibility that at least some lytic transcripts derived from these latency gene loci may have unique, non-coding nuclear functions during reactivation. Our analysis has also identified two previously unknown genes, BCLT1 and BCRT2, that map to the BamHI C-region of the EBV genome. Pathway analysis of cellular gene expression changes following BCR activation identified inflammatory response as the top predicted function, and ILK and TREM1 as the top predicted canonical pathways.