BamHI-A rightward frame 1, an Epstein-Barr virus-encoded oncogene and immune modulator

Abstract

Epstein-Barr virus (EBV) causes several benign and malignant disorders of lymphoid and epithelial origin. EBV-related tumors display distinct patterns of viral latent gene expression, of which the BamHI-A rightward frame 1 (BARF1) is selectively expressed in carcinomas, regulated by cellular differentiation factors including ΔNp63α. BARF1 functions as a viral oncogene, immortalizing and transforming epithelial cells of different origin by acting as a mitogenic growth factor, inducing cyclin-D expression, and up-regulating antiapoptotic Bcl-2, stimulating host cell growth and survival. In addition, secreted hexameric BARF1 has immune evasive properties, functionally corrupting macrophage colony stimulating factor, as supported by recent functional and structural data. Therefore, BARF1, an intracellular and secreted protein, not only has multiple pathogenic functions but also can function as a target for immune responses. Deciphering the role of BARF1 in EBV biology will contribute to novel diagnostic and treatment options for EBV-driven carcinomas. Herein, we discuss recent insights on the regulation of BARF1 expression and aspects of structure-function relating to its oncogenic and immune suppressive properties.

Figure 1

Putative functions of BamHI-A rightward frame 1. Schematic representation of functions ascribed to BamHI-A rightward frame 1

Figure 2

Mutations and homology domain of BamHI-A rightward frame 1(BARF1) protein. (A) Schematic representation of the BARF1 221 peptide. Left of the dotted line is the intracellular N-terminal part. Frequent amino acid mutations are depicted in black, rare mutations are depicted in gray. White bars; structural loops that interact with M-CSF, black asterisk; high mannose N-linked glycosylation at N95, white asterisk; predicted O-glycosylation site at T169, black ovals; C146 and C201 involved in folding and oligomerization. (B) BARF1 has sequence homology with a conserved domain found in several growth factor receptors. BARF1 aa146–158 is shown with homologous regions from the indicated receptors. Adapted from ,,

Figure 3

Cartoon of the soluble hexameric molecule BamH1-A rightward frame 1 (BARF1) hexameric structure.  (A) Top view of the BARF1 hexamer, the N-linked glycosylation is shown in a stick formation, (B) side view of the BARF1 hexamer

Figure 4

Schematic representation of BamHI-A rightward frame 1 (BARF1) transcriptional regulation. During lytic reactivation BARF1 is directly transactivated by the immediate early transcription factor R. Recent findings indicate that the differentiation marker ΔNp63α might be the cell type-specific transcription regulator enabling BARF1 transcription in latent in epithelial cells. The BARF1 promoter region is depicted up to −679 nucleotides relative to the BARF1 start-codon encoding methionine (ATG) start site and black vertical lines indicate methylation sites. Arrows point to the R binding sites in the BARF1 promoter , shaded bars indicate sites likely to be the most important for ΔNp63α BARF1 transactivation

Figure 5

BamHI-A rightward frame 1 mRNA is abundantly detectable in nasopharyngeal (NP) brushings. Epstein–Barr nuclear antigen 1 and BamHI-A rightward frame 1 RNA expression in NP brushings samples of NP carcinoma patients as determined by nucleic acid sequence-based amplification, in relation to Epstein–Barr virus DNA load. Shown is an autoradiogram of nucleic acid sequence-based amplification products hybridized with a radioactive-labeled internal oligonucleotide probe

Figure 6

Anchorage-independent growth in soft agar. Unpublished results from our own group demonstrate contact independent growth of BamHI-A rightward frame 1 expressing 293HEK cells

Figure 7

Soluble hexameric molecule BamH1-A rightward frame 1(sBARF1) is a macrophage colony stimulating factor (M-CSF) decoy receptor. (A) MTT proliferation test was used as readout for both increased cell numbers and level of differentiation. The inhibitory effect of sBARF1 on the monocyte to macrophage differentiation can be seen from day 3. The untreated cells showed increased viability, whereas the viability of sBARF1 treated macrophages remained the same in time. Adapted from . (B) sBARF1 negatively influences monocyte to macrophage differentiation in response to M-CSF, here shown are the differences in morphology and viability between normal monocyte derived macrophages (MoФ) and MoФ differentiated in the presence of sBARF1 (sBARF1 MoФ). Adapted from Hoebe et al. . (C) Structure of the BARF1/M-CSF complex as by Shim et al. . One sBARF1 hexamer can bind three human M-CSF dimers