The Viral Oncoprotein vFLIP Encoded by KSHV/HHV-8 Is Responsible for the Unique Transcriptional Signature and Phenotypic Features of Primary Effusion Lymphoma

Primary effusion lymphomas (PELs) are caused by the Kaposi's sarcoma herpesvirus (KSHV, also called human herpesvirus 8, HHV-8). PELs have a distinctive immunophenotype, lacking expression of most B cell antigens. In addition, PELs have a very distinct gene expression signature when compared to that of other lymphoma subtypes. While KSHV encodes almost 100 proteins, only a handful is expressed in latently infected PEL cells. One of these is vFLIP, which activates NF-kB and inhibits apoptosis and autophagy, and causes tumors in mice. In this study we evaluated the global effect of ectopic expression of vFLIP in a B cell lymphoma cell line, as well as that of suppression of vFLIP expression in a PEL cell line.

vFLIP expression was knocked down with siRNA in the PEL cell line BC-3. Namalwa cells were stably transfected with a tetracycline-inducible vFLIP, and vFLIP expression was induced with doxycycline for 48 hours. The Namalwa cell line was selected because it contains EBV with a type I latency pattern, also a feature of most PELs. We evaluated RNA extracted from these vFLIP+ and FLIP– cells in a PEL and Burkitt lymphoma background for gene expression array analysis using the Illumina Direct Hybridization Assay. Results were validated by immunohistochemistry and flow cytometry.

The vFLIP signature in a BL cell line was analyzed by gene set enrichment analysis (GSEA), which is an analytical method to determine how gene sets from other public studies compare to our signature. 1,445 public gene signatures were compared to a rank ordered list of genes that were altered by expression of vFLIP. The top match among these previous gene signatures, for both vFLIP upregulated and downregulated genes, was the PEL signature as compared to other lymphoma subtypes and normal B cell subsets (Kelin et al, Blood 101, 2003). Thus, expression of vFLIP in BL leads to a switch from a BL expression signature to that of PEL. Furthermore, expression of vFLIP in BL cells let to an almost complete downregulation of several B cell antigens, including surface Ig light chains, CD19, as well as CD10, in conjunction with upregulation of CD138 expression.

The unique phenotype of PEL has long been an intriguing feature of this disease. We have found that a single gene encoded by KSHV, vFLIP, can induce a transcriptional signature of PEL, as well as the phenotype characteristic of this disease.

No relevant conflicts of interest to declare.