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Using high-throughput RNA sequencing data from 50 common lymphoma cell culture models from the Cancer Cell Line Encyclopedia project, we performed an unbiased global interrogation for the presence of a panel of 740 viruses and strains known to infect human and other mammalian cells. This led to the findings of previously identified infections by Epstein-Barr virus (EBV), Kaposi's sarcoma herpesvirus (KSHV), and human T-lymphotropic virus type 1 (HTLV-1). In addition, we also found a previously unreported infection of one cell line (DEL) with a murine leukemia virus. High expression of murine leukemia virus (MuLV) transcripts was observed in DEL cells, and we identified four transcriptionally active integration sites, one being in the TNFRSF6B gene. We also found low levels of MuLV reads in a number of other cell lines and provided evidence suggesting cross-contamination during sequencing. Analysis of HTLV-1 integrations in two cell lines, HuT 102 and MJ, identified 14 and 66 transcriptionally active integration sites with potentially activating integrations in immune regulatory genes, including interleukin-15 (IL-15), IL-6ST, STAT5B, HIVEP1, and IL-9R. Although KSHV and EBV do not typically integrate into the genome, we investigated a previously identified integration of EBV into the BACH2 locus in Raji cells. This analysis identified a BACH2 disruption mechanism involving splice donor sequestration. Through viral gene expression analysis, we detected expression of stable intronic RNAs from the EBV BamHI W repeats that may be part of long transcripts spanning the repeat region. We also observed transcripts at the EBV vIL-10 locus exclusively in the Hodgkin's lymphoma cell line, Hs 611.T, the expression of which were uncoupled from other lytic genes. Assessment of the KSHV viral transcriptome in BCP-1 cells showed expression of the viral immune regulators, K2/vIL-6, K4/vIL-8-like vCCL1, and K5/E2-ubiquitin ligase 1 that was significantly higher than expression of the latency-associated nuclear antigen. Together, this investigation sheds light into the virus composition across these lymphoma model systems and provides insights into common viral mechanistic principles.

High-Throughput RNA Sequencing-Based Virome Analysis of 50 Lymphoma Cell Lines from the Cancer Cell Line Encyclopedia Project