Quantitative and Qualitative RNA-Seq-Based Evaluation of Epstein-Barr Virus Transcription in Type I Latency Burkitt's Lymphoma Cells

Abstract

RNA-seq provides a rich source of transcriptome information with high qualitative and quantitative value. Here, we provide a pipeline for Epstein-Barr virus (EBV) transcriptome analysis using RNA-seq and we apply it to two type I latency cell lines, Mutu I and Akata. This analysis revealed substantial average expression levels of many lytic genes in predominantly latent cell populations. The lytic transcripts BHLF1 and LF3 were expressed at levels greater than those for 98% of all cellular polyadenylated transcripts. Exon junction mapping accurately identified the Qp-derived EBNA1 splicing pattern, lytic gene splicing, and a complex splicing pattern within the BamHI A region.

Identification of New Viral Genes and Transcript Isoforms during Epstein-Barr Virus Reactivation using RNA-Seq

“…We have previously reported a pipeline for EBV transcriptome analysis in the context of the human transcriptome using RNASeq (11). The pipeline was applied to whole-cell RNA preps from the type I latency cell lines Mutu I and Akata to illustrate the quantitative and qualitative value of RNA-Seq in assessing viral transcription.…”

“…The development of these tools for the analysis of the type I EBV strain is important since it is a more common strain observed in tissue culture models and in vivo. For most of the EBV genome, reasonable alignment data can be obtained even when using sequencing data from a type I strain and aligning to a type II strain genome (11). Nevertheless, the capture of sequences from less well- and BCRT2 (C) and 5= RACE identification of start sites.…”

Whole-Genome Sequencing of the Akata and Mutu Epstein-Barr Virus Strains

“…9; note that, on the sense strand shown in the genome browser view, this is depicted as a T-to-C change). These editing events were also observed using RNA-seq data from latent Mutu cells (14), and the orientation of this transcript and the associated RNA-editing event were verified using strand-specific sequencing data from induced Akata cells (unpublished). Nevertheless, in addition to the high abundance of RNA editing of the antisense transcripts, we also detected A-to-I (G) editing at the primary transcripts (primicroRNA [pri-miRNA] and/or pre-miRNA) of ebv-miR-BART6 (position 139637 of the Akata genome and position 139956 of the Mutu genome) consistent with previous studies in other cell lines (23).…”

“…NC_007605) (these regions are indicated in Table S2 in the supplemental material). Lastly, the finalized Akata and Mutu EBV genomes were further validated by visualization of alignments from both latent and lytic Akata RNA-seq data and from latent Mutu RNA-seq data (13,14).…”

“…Akata and Mutu EBV transcriptome analysis was performed by aligning sequence reads from latent and lytic cytoplasmic and nuclear Akata RNA (reported previously [13]) and from latent Mutu RNA (reported previously [14]) to the assembled Akata or Mutu EBV genome using Novoalign. Aligned reads were visualized using the IGV browser.…”